From 6407d5b5d6cba31da11ab6641b1a71b937720c65 Mon Sep 17 00:00:00 2001
From: GitHub Actions <actions@github.com>
Date: Thu, 2 May 2024 18:15:26 +0000
Subject: [PATCH] site deploy

Auto-generated via {sandpaper}
Source  : b321a7f4d5d1bf2338e8f61f7e3fd4b74d457ebf
Branch  : md-outputs
Author  : GitHub Actions <actions@github.com>
Time    : 2024-05-02 18:13:01 +0000
Message : markdown source builds

Auto-generated via {sandpaper}
Source  : ad7e2280e7c053d7cedbb3d9c213b87fde480992
Branch  : main
Author  : Anthony Christidis <anthony-alexander_christidis@hms.harvard.edu>
Time    : 2024-05-02 17:21:02 +0000
Message : Update cell type annotation & multi-sample analyses episodes (update time, questions and objectives).
---
 aio.html                                      | 135 ++++++++++-------
 ...k-1_879a1babfa2cb9874c41088352e40217.RData | Bin 3059 -> 3063 bytes
 ...k-2_3167a19dc7b8bd71e74cd2f0dc6f0e2d.RData | Bin 3127 -> 3129 bytes
 cell_type_annotation.html                     |  26 +++-
 eda_qc.html                                   |  10 +-
 hca.html                                      |  62 ++++----
 instructor/aio.html                           | 139 +++++++++++-------
 instructor/cell_type_annotation.html          |  28 +++-
 instructor/eda_qc.html                        |  10 +-
 instructor/hca.html                           |  62 ++++----
 instructor/index.html                         |  23 ++-
 instructor/intro-sce.html                     |   2 +-
 instructor/large_data.html                    |  37 ++---
 instructor/multi-sample.html                  |  30 +++-
 intro-sce.html                                |   2 +-
 large_data.html                               |  37 ++---
 md5sum.txt                                    |   4 +-
 multi-sample.html                             |  28 +++-
 pkgdown.yml                                   |   2 +-
 19 files changed, 394 insertions(+), 243 deletions(-)

diff --git a/aio.html b/aio.html
index 9c912ab..2237d40 100644
--- a/aio.html
+++ b/aio.html
@@ -782,7 +782,7 @@ <h3 class="callout-title">Exercise 1<a class="anchor" aria-label="anchor" href="
 <button class="accordion-button hint-button collapsed" type="button" data-bs-toggle="collapse" data-bs-target="#collapseHint1" aria-expanded="false" aria-controls="collapseHint1">
   <h4 class="accordion-header" id="headingHint1">Give me a hint</h4>
 </button>
-<div id="collapseHint1" class="accordion-collapse collapse" data-bs-parent="#accordionHint1" aria-labelledby="headingHint1">
+<div id="collapseHint1" class="accordion-collapse collapse" aria-labelledby="headingHint1" data-bs-parent="#accordionHint1">
 <div class="accordion-body">
 <p>The <code>SingleCellExperiment</code> function can be used to create
 a new SingleCellExperiment object</p>
@@ -1030,7 +1030,7 @@ <h3 class="code-label">OUTPUT<i aria-hidden="true" data-feather="chevron-left"><
   <h3 class="accordion-header" id="headingSpoiler1">
 <div class="note-square"><i aria-hidden="true" class="callout-icon" data-feather="eye"></i></div>Setting the Random Seed</h3>
 </button>
-<div id="collapseSpoiler1" class="accordion-collapse collapse" aria-labelledby="headingSpoiler1" data-bs-parent="#accordionSpoiler1">
+<div id="collapseSpoiler1" class="accordion-collapse collapse" data-bs-parent="#accordionSpoiler1" aria-labelledby="headingSpoiler1">
 <div class="accordion-body">
 <p>Whenever your code involves randomness, it’s a good idea to set the
 random seed in R with <code>set.seed()</code> from the base <a href="https://stat.ethz.ch/R-manual/R-devel/library/base/html/Random.html" class="external-link">random</a>
@@ -1839,7 +1839,7 @@ <h3 class="callout-title">Exercise 2: Dimensionality Reduction<a class="anchor"
 <button class="accordion-button hint-button collapsed" type="button" data-bs-toggle="collapse" data-bs-target="#collapseHint1" aria-expanded="false" aria-controls="collapseHint1">
   <h4 class="accordion-header" id="headingHint1">Give me a hint</h4>
 </button>
-<div id="collapseHint1" class="accordion-collapse collapse" aria-labelledby="headingHint1" data-bs-parent="#accordionHint1">
+<div id="collapseHint1" class="accordion-collapse collapse" data-bs-parent="#accordionHint1" aria-labelledby="headingHint1">
 <div class="accordion-body">
 <p>First subset the object to include only highly variable genes
 (<code>sce2 &lt;- sce[hvg.sce.var,]</code>) and then apply the
@@ -1937,7 +1937,12 @@ <h2 class="card-header">Overview</h2>
 <div class="inner">
 <h3 class="card-title">Questions</h3>
 <ul>
-<li>TODO</li>
+<li>How is clustering used in scRNA-seq data analysis?</li>
+<li>How are marker genes used to interpret clustering results?</li>
+<li>What are the challenges and common approaches for assigning cell
+type labels in scRNA-seq data?</li>
+<li>How can we further validate and refine cell type annotations from
+scRNA-seq data?</li>
 </ul>
 </div>
 </div>
@@ -1947,7 +1952,18 @@ <h3 class="card-title">Questions</h3>
 <div class="inner bordered">
 <h3 class="card-title">Objectives</h3>
 <ul>
-<li>TODO</li>
+<li>Understand that clustering is an unsupervised learning technique
+that groups cells with similar expression profiles.</li>
+<li>Learn how to identify genes that differentiate clusters and assign
+potential biological meaning to each cluster based on marker gene
+function.</li>
+<li>Understand two strategies for cell type annotation: 1. comparing to
+reference datasets with known cell type labels and 2. Identifying cell
+type marker genes and assigning labels based on marker set
+enrichment.</li>
+<li>Explore additional methods to assess the quality of assigned labels,
+such as examining the distribution of marker gene expression within
+clusters.</li>
 </ul>
 </div>
 </div>
@@ -3152,7 +3168,16 @@ <h2 class="card-header">Overview</h2>
 <div class="inner">
 <h3 class="card-title">Questions</h3>
 <ul>
-<li>TODO</li>
+<li>How can we identify and remove unwanted variation introduced by
+processing differences between samples (batch effects) in scRNA-seq
+data?</li>
+<li>How can we identify genes that are expressed differently between
+groups of cells in scRNA-seq data (e.g., different cell types or
+conditions)?</li>
+<li>How can we identify cell types (clusters) that show significant
+changes in abundance between conditions in scRNA-seq data? How can we
+address potential biases caused by differences in cell numbers per
+type?</li>
 </ul>
 </div>
 </div>
@@ -3162,7 +3187,16 @@ <h3 class="card-title">Questions</h3>
 <div class="inner bordered">
 <h3 class="card-title">Objectives</h3>
 <ul>
-<li>TODO</li>
+<li>Understand the concept of batch effects and their impact on data
+analysis. Learn how to utilize tools like correctExperiment from the
+batchelor package to correct for batch effects based on sample
+information.</li>
+<li>Master the workflow for differential expression analysis (DEA),
+including grouping cells, creating pseudo-bulk samples, normalization,
+and differential expression testing with tools like edgeR.</li>
+<li>Understand differential abundance (DA) analysis, considering
+compositional effects and strategies to address them, for robust
+identification of cell type abundance changes between conditions.</li>
 </ul>
 </div>
 </div>
@@ -4651,21 +4685,22 @@ <h3 class="code-label">R<i aria-hidden="true" data-feather="chevron-left"></i><i
 <h3 class="code-label">OUTPUT<i aria-hidden="true" data-feather="chevron-left"></i><i aria-hidden="true" data-feather="chevron-right"></i>
 </h3>
 <pre class="output" tabindex="0"><code>     approx
-exact   1   2   3   4   5   6   7   8   9  10  11  12  13  14
-   1   88   0   0   0   2   0   0   0   2   0   0   0   0   0
-   2    0 143   0   1   0   0   0   0   0   0   0   0   0   0
-   3    0   0  75   0   2   0   0   0   0   0   0   0   0   0
-   4    0   0   0 397   0   0   0   0   0   0   0   0   0   0
-   5    0   0   0   0 391   0   0   0   0   1   0   0   0   0
-   6    0   0   0   0   0 202   0   0   0   0   0   0   0   0
-   7    0   0   0   0   0   8 245   0   0   1   0   0   0   0
-   8    0   0   0   0   1   0   0  95   0   0   0   0   0   0
-   9    1   0   0   0   1   0   0   0 106   0   0   0   0   0
-   10   0   0   0   0   0   0   0   0   0 113   0   0   0   0
-   11   0   0   0   0   0   0   0   0   0   0 153   0   0   0
-   12   0   0   0   0   4   0   0   0   0  10   0 203   0   0
-   13   0   0   0   0   0   0   0   0   0   0   0   0 146   0
-   14   0   0   0   0   0   0   0   0   0   0   0   0   0  20</code></pre>
+exact   1   2   3   4   5   6   7   8   9  10  11  12  13  14  15
+   1   90   0   0   0   0   0   0   0   1   0   0   0   0   0   0
+   2    0 143   0   0   0   0   0   0   0   0   0   0   0   0   1
+   3    0   0  75   0   0   0   0   0   0   0   0   0   0   0   0
+   4    0   0   0 341   0   0   0   0   0   0   0   0   0   0   0
+   5    0   0   2   0 391   0   0   2   0   0   0   5   0   0   0
+   6    0   0   0   0   0  80 245   0   0   1   0   2   0   0   0
+   7    0   0   0   0   0 129   0   0   0   0   2   0   0   0   0
+   8    0   0   0   0   1   0   0  93   0   0   0   0   0   0   0
+   9    1   0   0   0   1   0   0   0 106   0   0   0   0   0   0
+   10   0   0   0   0   0   0   0   0   0 108   8   5   0   0   0
+   11   0   0   0   0   0   0   0   0   0   0 138   0   0   0   0
+   12   0   0   0   0   5   0   0   0   0   0   0 207   0   0   0
+   13   0   0   0   0   0   0   0   0   0   0   6   0 146   0   0
+   14   0   0   0   0   0   0   0   0   0   0   0   0   0  20   0
+   15   0   0   0   0   0   0   0   0   0   0   0   0   0   0  56</code></pre>
 </div>
 <p>Note that Annoy writes the NN index to disk prior to performing the
 search. Thus, it may not actually be faster than the default exact
@@ -5482,18 +5517,18 @@ <h3 class="code-label">OUTPUT<i aria-hidden="true" data-feather="chevron-left"><
 </h3>
 <pre class="output" tabindex="0"><code># Source:   SQL [?? x 2]
 # Database: DuckDB v0.10.2 [unknown@Linux 6.5.0-1018-azure:R 4.3.3/:memory:]
-   tissue                             n
-   &lt;chr&gt;                          &lt;dbl&gt;
- 1 blood                             17
- 2 kidney                             8
- 3 kidney blood vessel                4
- 4 lung parenchyma                    2
- 5 respiratory airway                 6
- 6 dorsolateral prefrontal cortex     1
- 7 heart left ventricle               7
- 8 bone marrow                        4
- 9 lung                               4
-10 sigmoid colon                      1
+   tissue                      n
+   &lt;chr&gt;                   &lt;dbl&gt;
+ 1 pleural effusion            1
+ 2 caudate lobe of liver       1
+ 3 cortex of kidney            7
+ 4 nose                        1
+ 5 thymus                      4
+ 6 epithelium of esophagus     1
+ 7 transverse colon            1
+ 8 liver                       1
+ 9 renal medulla               6
+10 caecum                      1
 # ℹ more rows</code></pre>
 </div>
 </div>
@@ -5528,18 +5563,18 @@ <h3 class="code-label">OUTPUT<i aria-hidden="true" data-feather="chevron-left"><
 </h3>
 <pre class="output" tabindex="0"><code># Source:   SQL [?? x 2]
 # Database: DuckDB v0.10.2 [unknown@Linux 6.5.0-1018-azure:R 4.3.3/:memory:]
-   assay                              n
-   &lt;chr&gt;                          &lt;dbl&gt;
- 1 sci-RNA-seq                        1
- 2 10x 5' v2                          2
- 3 10x 3' v1                          1
- 4 Smart-seq2                         1
- 5 10x 3' v3                         21
- 6 Slide-seq                          4
- 7 10x 3' v2                         27
- 8 Visium Spatial Gene Expression     7
- 9 10x 5' v1                          7
-10 scRNA-seq                          4
+   assay           n
+   &lt;chr&gt;       &lt;dbl&gt;
+ 1 10x 3' v3      21
+ 2 Slide-seq       4
+ 3 scRNA-seq       4
+ 4 Seq-Well        2
+ 5 Drop-seq        1
+ 6 10x 5' v2       2
+ 7 10x 3' v1       1
+ 8 Smart-seq2      1
+ 9 sci-RNA-seq     1
+10 10x 3' v2      27
 # ℹ more rows</code></pre>
 </div>
 </div>
@@ -5743,7 +5778,7 @@ <h3 class="callout-title">Exercise 1<a class="anchor" aria-label="anchor" href="
 <button class="accordion-button solution-button collapsed" type="button" data-bs-toggle="collapse" data-bs-target="#collapseSolution1" aria-expanded="false" aria-controls="collapseSolution1">
   <h4 class="accordion-header" id="headingSolution1">Show me the solution</h4>
 </button>
-<div id="collapseSolution1" class="accordion-collapse collapse" aria-labelledby="headingSolution1" data-bs-parent="#accordionSolution1">
+<div id="collapseSolution1" class="accordion-collapse collapse" data-bs-parent="#accordionSolution1" aria-labelledby="headingSolution1">
 <div class="accordion-body">
 <div class="codewrapper sourceCode" id="cb24">
 <h3 class="code-label">R<i aria-hidden="true" data-feather="chevron-left"></i><i aria-hidden="true" data-feather="chevron-right"></i>
@@ -5777,7 +5812,7 @@ <h3 class="callout-title">Exercise 2<a class="anchor" aria-label="anchor" href="
 <button class="accordion-button solution-button collapsed" type="button" data-bs-toggle="collapse" data-bs-target="#collapseSolution2" aria-expanded="false" aria-controls="collapseSolution2">
   <h4 class="accordion-header" id="headingSolution2">Show me the solution</h4>
 </button>
-<div id="collapseSolution2" class="accordion-collapse collapse" aria-labelledby="headingSolution2" data-bs-parent="#accordionSolution2">
+<div id="collapseSolution2" class="accordion-collapse collapse" data-bs-parent="#accordionSolution2" aria-labelledby="headingSolution2">
 <div class="accordion-body">
 <div class="codewrapper sourceCode" id="cb25">
 <h3 class="code-label">R<i aria-hidden="true" data-feather="chevron-left"></i><i aria-hidden="true" data-feather="chevron-right"></i>
@@ -5811,7 +5846,7 @@ <h3 class="callout-title">Exercise 3<a class="anchor" aria-label="anchor" href="
 <button class="accordion-button solution-button collapsed" type="button" data-bs-toggle="collapse" data-bs-target="#collapseSolution3" aria-expanded="false" aria-controls="collapseSolution3">
   <h4 class="accordion-header" id="headingSolution3">Show me the solution</h4>
 </button>
-<div id="collapseSolution3" class="accordion-collapse collapse" aria-labelledby="headingSolution3" data-bs-parent="#accordionSolution3">
+<div id="collapseSolution3" class="accordion-collapse collapse" data-bs-parent="#accordionSolution3" aria-labelledby="headingSolution3">
 <div class="accordion-body">
 <div class="codewrapper sourceCode" id="cb26">
 <h3 class="code-label">R<i aria-hidden="true" data-feather="chevron-left"></i><i aria-hidden="true" data-feather="chevron-right"></i>
@@ -5853,7 +5888,7 @@ <h3 class="callout-title">Exercise 4<a class="anchor" aria-label="anchor" href="
 <button class="accordion-button solution-button collapsed" type="button" data-bs-toggle="collapse" data-bs-target="#collapseSolution4" aria-expanded="false" aria-controls="collapseSolution4">
   <h4 class="accordion-header" id="headingSolution4">Show me the solution</h4>
 </button>
-<div id="collapseSolution4" class="accordion-collapse collapse" aria-labelledby="headingSolution4" data-bs-parent="#accordionSolution4">
+<div id="collapseSolution4" class="accordion-collapse collapse" data-bs-parent="#accordionSolution4" aria-labelledby="headingSolution4">
 <div class="accordion-body">
 <div class="codewrapper sourceCode" id="cb27">
 <h3 class="code-label">R<i aria-hidden="true" data-feather="chevron-left"></i><i aria-hidden="true" data-feather="chevron-right"></i>
diff --git a/cache/unnamed-chunk-1_879a1babfa2cb9874c41088352e40217.RData b/cache/unnamed-chunk-1_879a1babfa2cb9874c41088352e40217.RData
index 1400234d67ba6b323f49536b57873f4757803445..ae68a2e1df74abc75674169fec48c9d35a7e3f63 100644
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diff --git a/cell_type_annotation.html b/cell_type_annotation.html
index e0b4b63..77c45f7 100644
--- a/cell_type_annotation.html
+++ b/cell_type_annotation.html
@@ -92,11 +92,11 @@
   Orchestrating Large-Scale Single-Cell Analysis with Bioconductor
 </div>
 
-<aside class="col-md-12 lesson-progress"><div style="width: 53%" class="percentage">
-    53%
+<aside class="col-md-12 lesson-progress"><div style="width: 36%" class="percentage">
+    36%
   </div>
   <div class="progress incubator">
-    <div class="progress-bar incubator" role="progressbar" style="width: 53%" aria-valuenow="53" aria-label="Lesson Progress" aria-valuemin="0" aria-valuemax="100">
+    <div class="progress-bar incubator" role="progressbar" style="width: 36%" aria-valuenow="36" aria-label="Lesson Progress" aria-valuemin="0" aria-valuemax="100">
     </div>
   </div>
 </aside><div class="container">
@@ -285,7 +285,12 @@ <h2 class="card-header">Overview</h2>
 <div class="card-body">
 <div class="inner">
 <h3 class="card-title">Questions</h3>
-<ul><li>TODO</li>
+<ul><li>How is clustering used in scRNA-seq data analysis?</li>
+<li>How are marker genes used to interpret clustering results?</li>
+<li>What are the challenges and common approaches for assigning cell
+type labels in scRNA-seq data?</li>
+<li>How can we further validate and refine cell type annotations from
+scRNA-seq data?</li>
 </ul></div>
 </div>
 </div>
@@ -293,7 +298,18 @@ <h3 class="card-title">Questions</h3>
 <div class="card-body">
 <div class="inner bordered">
 <h3 class="card-title">Objectives</h3>
-<ul><li>TODO</li>
+<ul><li>Understand that clustering is an unsupervised learning technique
+that groups cells with similar expression profiles.</li>
+<li>Learn how to identify genes that differentiate clusters and assign
+potential biological meaning to each cluster based on marker gene
+function.</li>
+<li>Understand two strategies for cell type annotation: 1. comparing to
+reference datasets with known cell type labels and 2. Identifying cell
+type marker genes and assigning labels based on marker set
+enrichment.</li>
+<li>Explore additional methods to assess the quality of assigned labels,
+such as examining the distribution of marker gene expression within
+clusters.</li>
 </ul></div>
 </div>
 </div>
diff --git a/eda_qc.html b/eda_qc.html
index 400afbf..1c927a2 100644
--- a/eda_qc.html
+++ b/eda_qc.html
@@ -92,11 +92,11 @@
   Orchestrating Large-Scale Single-Cell Analysis with Bioconductor
 </div>
 
-<aside class="col-md-12 lesson-progress"><div style="width: 21%" class="percentage">
-    21%
+<aside class="col-md-12 lesson-progress"><div style="width: 14%" class="percentage">
+    14%
   </div>
   <div class="progress incubator">
-    <div class="progress-bar incubator" role="progressbar" style="width: 21%" aria-valuenow="21" aria-label="Lesson Progress" aria-valuemin="0" aria-valuemax="100">
+    <div class="progress-bar incubator" role="progressbar" style="width: 14%" aria-valuenow="14" aria-label="Lesson Progress" aria-valuemin="0" aria-valuemax="100">
     </div>
   </div>
 </aside><div class="container">
@@ -459,7 +459,7 @@ <h3 class="code-label">OUTPUT<i aria-hidden="true" data-feather="chevron-left"><
   <h3 class="accordion-header" id="headingSpoiler1">
 <div class="note-square"><i aria-hidden="true" class="callout-icon" data-feather="eye"></i></div>Setting the Random Seed</h3>
 </button>
-<div id="collapseSpoiler1" class="accordion-collapse collapse" aria-labelledby="headingSpoiler1" data-bs-parent="#accordionSpoiler1">
+<div id="collapseSpoiler1" class="accordion-collapse collapse" data-bs-parent="#accordionSpoiler1" aria-labelledby="headingSpoiler1">
 <div class="accordion-body">
 <p>Whenever your code involves randomness, it’s a good idea to set the
 random seed in R with <code>set.seed()</code> from the base <a href="https://stat.ethz.ch/R-manual/R-devel/library/base/html/Random.html" class="external-link">random</a>
@@ -1247,7 +1247,7 @@ <h3 class="callout-title">Exercise 2: Dimensionality Reduction<a class="anchor"
 <button class="accordion-button hint-button collapsed" type="button" data-bs-toggle="collapse" data-bs-target="#collapseHint1" aria-expanded="false" aria-controls="collapseHint1">
   <h4 class="accordion-header" id="headingHint1">Give me a hint</h4>
 </button>
-<div id="collapseHint1" class="accordion-collapse collapse" aria-labelledby="headingHint1" data-bs-parent="#accordionHint1">
+<div id="collapseHint1" class="accordion-collapse collapse" data-bs-parent="#accordionHint1" aria-labelledby="headingHint1">
 <div class="accordion-body">
 <p>First subset the object to include only highly variable genes
 (<code>sce2 &lt;- sce[hvg.sce.var,]</code>) and then apply the
diff --git a/hca.html b/hca.html
index 0cc21f7..6f0fdeb 100644
--- a/hca.html
+++ b/hca.html
@@ -92,11 +92,11 @@
   Orchestrating Large-Scale Single-Cell Analysis with Bioconductor
 </div>
 
-<aside class="col-md-12 lesson-progress"><div style="width: 79%" class="percentage">
-    79%
+<aside class="col-md-12 lesson-progress"><div style="width: 86%" class="percentage">
+    86%
   </div>
   <div class="progress incubator">
-    <div class="progress-bar incubator" role="progressbar" style="width: 79%" aria-valuenow="79" aria-label="Lesson Progress" aria-valuemin="0" aria-valuemax="100">
+    <div class="progress-bar incubator" role="progressbar" style="width: 86%" aria-valuenow="86" aria-label="Lesson Progress" aria-valuemin="0" aria-valuemax="100">
     </div>
   </div>
 </aside><div class="container">
@@ -488,18 +488,18 @@ <h3 class="code-label">OUTPUT<i aria-hidden="true" data-feather="chevron-left"><
 </h3>
 <pre class="output" tabindex="0"><code># Source:   SQL [?? x 2]
 # Database: DuckDB v0.10.2 [unknown@Linux 6.5.0-1018-azure:R 4.3.3/:memory:]
-   tissue                             n
-   &lt;chr&gt;                          &lt;dbl&gt;
- 1 blood                             17
- 2 kidney                             8
- 3 kidney blood vessel                4
- 4 lung parenchyma                    2
- 5 respiratory airway                 6
- 6 dorsolateral prefrontal cortex     1
- 7 heart left ventricle               7
- 8 bone marrow                        4
- 9 lung                               4
-10 sigmoid colon                      1
+   tissue                      n
+   &lt;chr&gt;                   &lt;dbl&gt;
+ 1 pleural effusion            1
+ 2 caudate lobe of liver       1
+ 3 cortex of kidney            7
+ 4 nose                        1
+ 5 thymus                      4
+ 6 epithelium of esophagus     1
+ 7 transverse colon            1
+ 8 liver                       1
+ 9 renal medulla               6
+10 caecum                      1
 # ℹ more rows</code></pre>
 </div>
 </div>
@@ -532,18 +532,18 @@ <h3 class="code-label">OUTPUT<i aria-hidden="true" data-feather="chevron-left"><
 </h3>
 <pre class="output" tabindex="0"><code># Source:   SQL [?? x 2]
 # Database: DuckDB v0.10.2 [unknown@Linux 6.5.0-1018-azure:R 4.3.3/:memory:]
-   assay                              n
-   &lt;chr&gt;                          &lt;dbl&gt;
- 1 sci-RNA-seq                        1
- 2 10x 5' v2                          2
- 3 10x 3' v1                          1
- 4 Smart-seq2                         1
- 5 10x 3' v3                         21
- 6 Slide-seq                          4
- 7 10x 3' v2                         27
- 8 Visium Spatial Gene Expression     7
- 9 10x 5' v1                          7
-10 scRNA-seq                          4
+   assay           n
+   &lt;chr&gt;       &lt;dbl&gt;
+ 1 10x 3' v3      21
+ 2 Slide-seq       4
+ 3 scRNA-seq       4
+ 4 Seq-Well        2
+ 5 Drop-seq        1
+ 6 10x 5' v2       2
+ 7 10x 3' v1       1
+ 8 Smart-seq2      1
+ 9 sci-RNA-seq     1
+10 10x 3' v2      27
 # ℹ more rows</code></pre>
 </div>
 </div>
@@ -738,7 +738,7 @@ <h3 class="callout-title">Exercise 1<a class="anchor" aria-label="anchor" href="
 <button class="accordion-button solution-button collapsed" type="button" data-bs-toggle="collapse" data-bs-target="#collapseSolution1" aria-expanded="false" aria-controls="collapseSolution1">
   <h4 class="accordion-header" id="headingSolution1">Show me the solution</h4>
 </button>
-<div id="collapseSolution1" class="accordion-collapse collapse" aria-labelledby="headingSolution1" data-bs-parent="#accordionSolution1">
+<div id="collapseSolution1" class="accordion-collapse collapse" data-bs-parent="#accordionSolution1" aria-labelledby="headingSolution1">
 <div class="accordion-body">
 <div class="codewrapper sourceCode" id="cb24">
 <h3 class="code-label">R<i aria-hidden="true" data-feather="chevron-left"></i><i aria-hidden="true" data-feather="chevron-right"></i>
@@ -772,7 +772,7 @@ <h3 class="callout-title">Exercise 2<a class="anchor" aria-label="anchor" href="
 <button class="accordion-button solution-button collapsed" type="button" data-bs-toggle="collapse" data-bs-target="#collapseSolution2" aria-expanded="false" aria-controls="collapseSolution2">
   <h4 class="accordion-header" id="headingSolution2">Show me the solution</h4>
 </button>
-<div id="collapseSolution2" class="accordion-collapse collapse" aria-labelledby="headingSolution2" data-bs-parent="#accordionSolution2">
+<div id="collapseSolution2" class="accordion-collapse collapse" data-bs-parent="#accordionSolution2" aria-labelledby="headingSolution2">
 <div class="accordion-body">
 <div class="codewrapper sourceCode" id="cb25">
 <h3 class="code-label">R<i aria-hidden="true" data-feather="chevron-left"></i><i aria-hidden="true" data-feather="chevron-right"></i>
@@ -806,7 +806,7 @@ <h3 class="callout-title">Exercise 3<a class="anchor" aria-label="anchor" href="
 <button class="accordion-button solution-button collapsed" type="button" data-bs-toggle="collapse" data-bs-target="#collapseSolution3" aria-expanded="false" aria-controls="collapseSolution3">
   <h4 class="accordion-header" id="headingSolution3">Show me the solution</h4>
 </button>
-<div id="collapseSolution3" class="accordion-collapse collapse" aria-labelledby="headingSolution3" data-bs-parent="#accordionSolution3">
+<div id="collapseSolution3" class="accordion-collapse collapse" data-bs-parent="#accordionSolution3" aria-labelledby="headingSolution3">
 <div class="accordion-body">
 <div class="codewrapper sourceCode" id="cb26">
 <h3 class="code-label">R<i aria-hidden="true" data-feather="chevron-left"></i><i aria-hidden="true" data-feather="chevron-right"></i>
@@ -848,7 +848,7 @@ <h3 class="callout-title">Exercise 4<a class="anchor" aria-label="anchor" href="
 <button class="accordion-button solution-button collapsed" type="button" data-bs-toggle="collapse" data-bs-target="#collapseSolution4" aria-expanded="false" aria-controls="collapseSolution4">
   <h4 class="accordion-header" id="headingSolution4">Show me the solution</h4>
 </button>
-<div id="collapseSolution4" class="accordion-collapse collapse" aria-labelledby="headingSolution4" data-bs-parent="#accordionSolution4">
+<div id="collapseSolution4" class="accordion-collapse collapse" data-bs-parent="#accordionSolution4" aria-labelledby="headingSolution4">
 <div class="accordion-body">
 <div class="codewrapper sourceCode" id="cb27">
 <h3 class="code-label">R<i aria-hidden="true" data-feather="chevron-left"></i><i aria-hidden="true" data-feather="chevron-right"></i>
diff --git a/instructor/aio.html b/instructor/aio.html
index 8791b93..fc45de7 100644
--- a/instructor/aio.html
+++ b/instructor/aio.html
@@ -785,7 +785,7 @@ <h3 class="callout-title">Exercise 1<a class="anchor" aria-label="anchor" href="
 <button class="accordion-button hint-button collapsed" type="button" data-bs-toggle="collapse" data-bs-target="#collapseHint1" aria-expanded="false" aria-controls="collapseHint1">
   <h4 class="accordion-header" id="headingHint1">Give me a hint</h4>
 </button>
-<div id="collapseHint1" class="accordion-collapse collapse" data-bs-parent="#accordionHint1" aria-labelledby="headingHint1">
+<div id="collapseHint1" class="accordion-collapse collapse" aria-labelledby="headingHint1" data-bs-parent="#accordionHint1">
 <div class="accordion-body">
 <p>The <code>SingleCellExperiment</code> function can be used to create
 a new SingleCellExperiment object</p>
@@ -1034,7 +1034,7 @@ <h3 class="code-label">OUTPUT<i aria-hidden="true" data-feather="chevron-left"><
   <h3 class="accordion-header" id="headingSpoiler1">
 <div class="note-square"><i aria-hidden="true" class="callout-icon" data-feather="eye"></i></div>Setting the Random Seed</h3>
 </button>
-<div id="collapseSpoiler1" class="accordion-collapse collapse" aria-labelledby="headingSpoiler1" data-bs-parent="#accordionSpoiler1">
+<div id="collapseSpoiler1" class="accordion-collapse collapse" data-bs-parent="#accordionSpoiler1" aria-labelledby="headingSpoiler1">
 <div class="accordion-body">
 <p>Whenever your code involves randomness, it’s a good idea to set the
 random seed in R with <code>set.seed()</code> from the base <a href="https://stat.ethz.ch/R-manual/R-devel/library/base/html/Random.html" class="external-link">random</a>
@@ -1843,7 +1843,7 @@ <h3 class="callout-title">Exercise 2: Dimensionality Reduction<a class="anchor"
 <button class="accordion-button hint-button collapsed" type="button" data-bs-toggle="collapse" data-bs-target="#collapseHint1" aria-expanded="false" aria-controls="collapseHint1">
   <h4 class="accordion-header" id="headingHint1">Give me a hint</h4>
 </button>
-<div id="collapseHint1" class="accordion-collapse collapse" aria-labelledby="headingHint1" data-bs-parent="#accordionHint1">
+<div id="collapseHint1" class="accordion-collapse collapse" data-bs-parent="#accordionHint1" aria-labelledby="headingHint1">
 <div class="accordion-body">
 <p>First subset the object to include only highly variable genes
 (<code>sce2 &lt;- sce[hvg.sce.var,]</code>) and then apply the
@@ -1930,7 +1930,7 @@ <h3 class="callout-title">Further Reading<a class="anchor" aria-label="anchor" h
 <p>Last updated on 2024-05-02 |
         
         <a href="https://github.com/ccb-hms/osca-carpentries/edit/main/episodes/cell_type_annotation.Rmd" class="external-link">Edit this page <i aria-hidden="true" data-feather="edit"></i></a></p>
-<p>Estimated time: <i aria-hidden="true" data-feather="clock"></i> 12 minutes</p>
+<p>Estimated time: <i aria-hidden="true" data-feather="clock"></i> 45 minutes</p>
 <div class="text-end">
           <button role="button" aria-pressed="false" tabindex="0" id="expand-code" class="pull-right" data-expand="Expand All Solutions " data-collapse="Collapse All Solutions "> Expand All Solutions <i aria-hidden="true" data-feather="plus"></i></button>
         </div>
@@ -1942,7 +1942,12 @@ <h2 class="card-header">Overview</h2>
 <div class="inner">
 <h3 class="card-title">Questions</h3>
 <ul>
-<li>TODO</li>
+<li>How is clustering used in scRNA-seq data analysis?</li>
+<li>How are marker genes used to interpret clustering results?</li>
+<li>What are the challenges and common approaches for assigning cell
+type labels in scRNA-seq data?</li>
+<li>How can we further validate and refine cell type annotations from
+scRNA-seq data?</li>
 </ul>
 </div>
 </div>
@@ -1952,7 +1957,18 @@ <h3 class="card-title">Questions</h3>
 <div class="inner bordered">
 <h3 class="card-title">Objectives</h3>
 <ul>
-<li>TODO</li>
+<li>Understand that clustering is an unsupervised learning technique
+that groups cells with similar expression profiles.</li>
+<li>Learn how to identify genes that differentiate clusters and assign
+potential biological meaning to each cluster based on marker gene
+function.</li>
+<li>Understand two strategies for cell type annotation: 1. comparing to
+reference datasets with known cell type labels and 2. Identifying cell
+type marker genes and assigning labels based on marker set
+enrichment.</li>
+<li>Explore additional methods to assess the quality of assigned labels,
+such as examining the distribution of marker gene expression within
+clusters.</li>
 </ul>
 </div>
 </div>
@@ -3146,7 +3162,7 @@ <h3 class="callout-title">Key Points<a class="anchor" aria-label="anchor" href="
 <p>Last updated on 2024-05-02 |
         
         <a href="https://github.com/ccb-hms/osca-carpentries/edit/main/episodes/multi-sample.Rmd" class="external-link">Edit this page <i aria-hidden="true" data-feather="edit"></i></a></p>
-<p>Estimated time: <i aria-hidden="true" data-feather="clock"></i> 12 minutes</p>
+<p>Estimated time: <i aria-hidden="true" data-feather="clock"></i> 45 minutes</p>
 <div class="text-end">
           <button role="button" aria-pressed="false" tabindex="0" id="expand-code" class="pull-right" data-expand="Expand All Solutions " data-collapse="Collapse All Solutions "> Expand All Solutions <i aria-hidden="true" data-feather="plus"></i></button>
         </div>
@@ -3158,7 +3174,16 @@ <h2 class="card-header">Overview</h2>
 <div class="inner">
 <h3 class="card-title">Questions</h3>
 <ul>
-<li>TODO</li>
+<li>How can we identify and remove unwanted variation introduced by
+processing differences between samples (batch effects) in scRNA-seq
+data?</li>
+<li>How can we identify genes that are expressed differently between
+groups of cells in scRNA-seq data (e.g., different cell types or
+conditions)?</li>
+<li>How can we identify cell types (clusters) that show significant
+changes in abundance between conditions in scRNA-seq data? How can we
+address potential biases caused by differences in cell numbers per
+type?</li>
 </ul>
 </div>
 </div>
@@ -3168,7 +3193,16 @@ <h3 class="card-title">Questions</h3>
 <div class="inner bordered">
 <h3 class="card-title">Objectives</h3>
 <ul>
-<li>TODO</li>
+<li>Understand the concept of batch effects and their impact on data
+analysis. Learn how to utilize tools like correctExperiment from the
+batchelor package to correct for batch effects based on sample
+information.</li>
+<li>Master the workflow for differential expression analysis (DEA),
+including grouping cells, creating pseudo-bulk samples, normalization,
+and differential expression testing with tools like edgeR.</li>
+<li>Understand differential abundance (DA) analysis, considering
+compositional effects and strategies to address them, for robust
+identification of cell type abundance changes between conditions.</li>
 </ul>
 </div>
 </div>
@@ -4658,21 +4692,22 @@ <h3 class="code-label">R<i aria-hidden="true" data-feather="chevron-left"></i><i
 <h3 class="code-label">OUTPUT<i aria-hidden="true" data-feather="chevron-left"></i><i aria-hidden="true" data-feather="chevron-right"></i>
 </h3>
 <pre class="output" tabindex="0"><code>     approx
-exact   1   2   3   4   5   6   7   8   9  10  11  12  13  14
-   1   88   0   0   0   2   0   0   0   2   0   0   0   0   0
-   2    0 143   0   1   0   0   0   0   0   0   0   0   0   0
-   3    0   0  75   0   2   0   0   0   0   0   0   0   0   0
-   4    0   0   0 397   0   0   0   0   0   0   0   0   0   0
-   5    0   0   0   0 391   0   0   0   0   1   0   0   0   0
-   6    0   0   0   0   0 202   0   0   0   0   0   0   0   0
-   7    0   0   0   0   0   8 245   0   0   1   0   0   0   0
-   8    0   0   0   0   1   0   0  95   0   0   0   0   0   0
-   9    1   0   0   0   1   0   0   0 106   0   0   0   0   0
-   10   0   0   0   0   0   0   0   0   0 113   0   0   0   0
-   11   0   0   0   0   0   0   0   0   0   0 153   0   0   0
-   12   0   0   0   0   4   0   0   0   0  10   0 203   0   0
-   13   0   0   0   0   0   0   0   0   0   0   0   0 146   0
-   14   0   0   0   0   0   0   0   0   0   0   0   0   0  20</code></pre>
+exact   1   2   3   4   5   6   7   8   9  10  11  12  13  14  15
+   1   90   0   0   0   0   0   0   0   1   0   0   0   0   0   0
+   2    0 143   0   0   0   0   0   0   0   0   0   0   0   0   1
+   3    0   0  75   0   0   0   0   0   0   0   0   0   0   0   0
+   4    0   0   0 341   0   0   0   0   0   0   0   0   0   0   0
+   5    0   0   2   0 391   0   0   2   0   0   0   5   0   0   0
+   6    0   0   0   0   0  80 245   0   0   1   0   2   0   0   0
+   7    0   0   0   0   0 129   0   0   0   0   2   0   0   0   0
+   8    0   0   0   0   1   0   0  93   0   0   0   0   0   0   0
+   9    1   0   0   0   1   0   0   0 106   0   0   0   0   0   0
+   10   0   0   0   0   0   0   0   0   0 108   8   5   0   0   0
+   11   0   0   0   0   0   0   0   0   0   0 138   0   0   0   0
+   12   0   0   0   0   5   0   0   0   0   0   0 207   0   0   0
+   13   0   0   0   0   0   0   0   0   0   0   6   0 146   0   0
+   14   0   0   0   0   0   0   0   0   0   0   0   0   0  20   0
+   15   0   0   0   0   0   0   0   0   0   0   0   0   0   0  56</code></pre>
 </div>
 <p>Note that Annoy writes the NN index to disk prior to performing the
 search. Thus, it may not actually be faster than the default exact
@@ -5490,18 +5525,18 @@ <h3 class="code-label">OUTPUT<i aria-hidden="true" data-feather="chevron-left"><
 </h3>
 <pre class="output" tabindex="0"><code># Source:   SQL [?? x 2]
 # Database: DuckDB v0.10.2 [unknown@Linux 6.5.0-1018-azure:R 4.3.3/:memory:]
-   tissue                             n
-   &lt;chr&gt;                          &lt;dbl&gt;
- 1 blood                             17
- 2 kidney                             8
- 3 kidney blood vessel                4
- 4 lung parenchyma                    2
- 5 respiratory airway                 6
- 6 dorsolateral prefrontal cortex     1
- 7 heart left ventricle               7
- 8 bone marrow                        4
- 9 lung                               4
-10 sigmoid colon                      1
+   tissue                      n
+   &lt;chr&gt;                   &lt;dbl&gt;
+ 1 pleural effusion            1
+ 2 caudate lobe of liver       1
+ 3 cortex of kidney            7
+ 4 nose                        1
+ 5 thymus                      4
+ 6 epithelium of esophagus     1
+ 7 transverse colon            1
+ 8 liver                       1
+ 9 renal medulla               6
+10 caecum                      1
 # ℹ more rows</code></pre>
 </div>
 </div>
@@ -5536,18 +5571,18 @@ <h3 class="code-label">OUTPUT<i aria-hidden="true" data-feather="chevron-left"><
 </h3>
 <pre class="output" tabindex="0"><code># Source:   SQL [?? x 2]
 # Database: DuckDB v0.10.2 [unknown@Linux 6.5.0-1018-azure:R 4.3.3/:memory:]
-   assay                              n
-   &lt;chr&gt;                          &lt;dbl&gt;
- 1 sci-RNA-seq                        1
- 2 10x 5' v2                          2
- 3 10x 3' v1                          1
- 4 Smart-seq2                         1
- 5 10x 3' v3                         21
- 6 Slide-seq                          4
- 7 10x 3' v2                         27
- 8 Visium Spatial Gene Expression     7
- 9 10x 5' v1                          7
-10 scRNA-seq                          4
+   assay           n
+   &lt;chr&gt;       &lt;dbl&gt;
+ 1 10x 3' v3      21
+ 2 Slide-seq       4
+ 3 scRNA-seq       4
+ 4 Seq-Well        2
+ 5 Drop-seq        1
+ 6 10x 5' v2       2
+ 7 10x 3' v1       1
+ 8 Smart-seq2      1
+ 9 sci-RNA-seq     1
+10 10x 3' v2      27
 # ℹ more rows</code></pre>
 </div>
 </div>
@@ -5751,7 +5786,7 @@ <h3 class="callout-title">Exercise 1<a class="anchor" aria-label="anchor" href="
 <button class="accordion-button solution-button collapsed" type="button" data-bs-toggle="collapse" data-bs-target="#collapseSolution1" aria-expanded="false" aria-controls="collapseSolution1">
   <h4 class="accordion-header" id="headingSolution1">Show me the solution</h4>
 </button>
-<div id="collapseSolution1" class="accordion-collapse collapse" aria-labelledby="headingSolution1" data-bs-parent="#accordionSolution1">
+<div id="collapseSolution1" class="accordion-collapse collapse" data-bs-parent="#accordionSolution1" aria-labelledby="headingSolution1">
 <div class="accordion-body">
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 <button class="accordion-button solution-button collapsed" type="button" data-bs-toggle="collapse" data-bs-target="#collapseSolution2" aria-expanded="false" aria-controls="collapseSolution2">
   <h4 class="accordion-header" id="headingSolution2">Show me the solution</h4>
 </button>
-<div id="collapseSolution2" class="accordion-collapse collapse" aria-labelledby="headingSolution2" data-bs-parent="#accordionSolution2">
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   <h4 class="accordion-header" id="headingSolution3">Show me the solution</h4>
 </button>
-<div id="collapseSolution3" class="accordion-collapse collapse" aria-labelledby="headingSolution3" data-bs-parent="#accordionSolution3">
+<div id="collapseSolution3" class="accordion-collapse collapse" data-bs-parent="#accordionSolution3" aria-labelledby="headingSolution3">
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@@ -5861,7 +5896,7 @@ <h3 class="callout-title">Exercise 4<a class="anchor" aria-label="anchor" href="
 <button class="accordion-button solution-button collapsed" type="button" data-bs-toggle="collapse" data-bs-target="#collapseSolution4" aria-expanded="false" aria-controls="collapseSolution4">
   <h4 class="accordion-header" id="headingSolution4">Show me the solution</h4>
 </button>
-<div id="collapseSolution4" class="accordion-collapse collapse" aria-labelledby="headingSolution4" data-bs-parent="#accordionSolution4">
+<div id="collapseSolution4" class="accordion-collapse collapse" data-bs-parent="#accordionSolution4" aria-labelledby="headingSolution4">
 <div class="accordion-body">
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diff --git a/instructor/cell_type_annotation.html b/instructor/cell_type_annotation.html
index d8633b4..7263120 100644
--- a/instructor/cell_type_annotation.html
+++ b/instructor/cell_type_annotation.html
@@ -92,11 +92,11 @@
   Orchestrating Large-Scale Single-Cell Analysis with Bioconductor
 </div>
 
-<aside class="col-md-12 lesson-progress"><div style="width: 53%" class="percentage">
-    53%
+<aside class="col-md-12 lesson-progress"><div style="width: 36%" class="percentage">
+    36%
   </div>
   <div class="progress incubator">
-    <div class="progress-bar incubator" role="progressbar" style="width: 53%" aria-valuenow="53" aria-label="Lesson Progress" aria-valuemin="0" aria-valuemax="100">
+    <div class="progress-bar incubator" role="progressbar" style="width: 36%" aria-valuenow="36" aria-label="Lesson Progress" aria-valuemin="0" aria-valuemax="100">
     </div>
   </div>
 </aside><div class="container">
@@ -272,7 +272,7 @@ <h1>Cell type annotation</h1>
         
         
         
-        <p>Estimated time: <i aria-hidden="true" data-feather="clock"></i> 12 minutes</p>
+        <p>Estimated time: <i aria-hidden="true" data-feather="clock"></i> 45 minutes</p>
         
         <div class="text-end">
           <button role="button" aria-pressed="false" tabindex="0" id="expand-code" class="pull-right" data-expand="Expand All Solutions " data-collapse="Collapse All Solutions "> Expand All Solutions <i aria-hidden="true" data-feather="plus"></i></button>
@@ -287,7 +287,12 @@ <h2 class="card-header">Overview</h2>
 <div class="card-body">
 <div class="inner">
 <h3 class="card-title">Questions</h3>
-<ul><li>TODO</li>
+<ul><li>How is clustering used in scRNA-seq data analysis?</li>
+<li>How are marker genes used to interpret clustering results?</li>
+<li>What are the challenges and common approaches for assigning cell
+type labels in scRNA-seq data?</li>
+<li>How can we further validate and refine cell type annotations from
+scRNA-seq data?</li>
 </ul></div>
 </div>
 </div>
@@ -295,7 +300,18 @@ <h3 class="card-title">Questions</h3>
 <div class="card-body">
 <div class="inner bordered">
 <h3 class="card-title">Objectives</h3>
-<ul><li>TODO</li>
+<ul><li>Understand that clustering is an unsupervised learning technique
+that groups cells with similar expression profiles.</li>
+<li>Learn how to identify genes that differentiate clusters and assign
+potential biological meaning to each cluster based on marker gene
+function.</li>
+<li>Understand two strategies for cell type annotation: 1. comparing to
+reference datasets with known cell type labels and 2. Identifying cell
+type marker genes and assigning labels based on marker set
+enrichment.</li>
+<li>Explore additional methods to assess the quality of assigned labels,
+such as examining the distribution of marker gene expression within
+clusters.</li>
 </ul></div>
 </div>
 </div>
diff --git a/instructor/eda_qc.html b/instructor/eda_qc.html
index f364e39..03d5f69 100644
--- a/instructor/eda_qc.html
+++ b/instructor/eda_qc.html
@@ -92,11 +92,11 @@
   Orchestrating Large-Scale Single-Cell Analysis with Bioconductor
 </div>
 
-<aside class="col-md-12 lesson-progress"><div style="width: 21%" class="percentage">
-    21%
+<aside class="col-md-12 lesson-progress"><div style="width: 14%" class="percentage">
+    14%
   </div>
   <div class="progress incubator">
-    <div class="progress-bar incubator" role="progressbar" style="width: 21%" aria-valuenow="21" aria-label="Lesson Progress" aria-valuemin="0" aria-valuemax="100">
+    <div class="progress-bar incubator" role="progressbar" style="width: 14%" aria-valuenow="14" aria-label="Lesson Progress" aria-valuemin="0" aria-valuemax="100">
     </div>
   </div>
 </aside><div class="container">
@@ -461,7 +461,7 @@ <h3 class="code-label">OUTPUT<i aria-hidden="true" data-feather="chevron-left"><
   <h3 class="accordion-header" id="headingSpoiler1">
 <div class="note-square"><i aria-hidden="true" class="callout-icon" data-feather="eye"></i></div>Setting the Random Seed</h3>
 </button>
-<div id="collapseSpoiler1" class="accordion-collapse collapse" aria-labelledby="headingSpoiler1" data-bs-parent="#accordionSpoiler1">
+<div id="collapseSpoiler1" class="accordion-collapse collapse" data-bs-parent="#accordionSpoiler1" aria-labelledby="headingSpoiler1">
 <div class="accordion-body">
 <p>Whenever your code involves randomness, it’s a good idea to set the
 random seed in R with <code>set.seed()</code> from the base <a href="https://stat.ethz.ch/R-manual/R-devel/library/base/html/Random.html" class="external-link">random</a>
@@ -1249,7 +1249,7 @@ <h3 class="callout-title">Exercise 2: Dimensionality Reduction<a class="anchor"
 <button class="accordion-button hint-button collapsed" type="button" data-bs-toggle="collapse" data-bs-target="#collapseHint1" aria-expanded="false" aria-controls="collapseHint1">
   <h4 class="accordion-header" id="headingHint1">Give me a hint</h4>
 </button>
-<div id="collapseHint1" class="accordion-collapse collapse" aria-labelledby="headingHint1" data-bs-parent="#accordionHint1">
+<div id="collapseHint1" class="accordion-collapse collapse" data-bs-parent="#accordionHint1" aria-labelledby="headingHint1">
 <div class="accordion-body">
 <p>First subset the object to include only highly variable genes
 (<code>sce2 &lt;- sce[hvg.sce.var,]</code>) and then apply the
diff --git a/instructor/hca.html b/instructor/hca.html
index 5a7ef51..956a050 100644
--- a/instructor/hca.html
+++ b/instructor/hca.html
@@ -92,11 +92,11 @@
   Orchestrating Large-Scale Single-Cell Analysis with Bioconductor
 </div>
 
-<aside class="col-md-12 lesson-progress"><div style="width: 79%" class="percentage">
-    79%
+<aside class="col-md-12 lesson-progress"><div style="width: 86%" class="percentage">
+    86%
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+    <div class="progress-bar incubator" role="progressbar" style="width: 86%" aria-valuenow="86" aria-label="Lesson Progress" aria-valuemin="0" aria-valuemax="100">
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@@ -490,18 +490,18 @@ <h3 class="code-label">OUTPUT<i aria-hidden="true" data-feather="chevron-left"><
 </h3>
 <pre class="output" tabindex="0"><code># Source:   SQL [?? x 2]
 # Database: DuckDB v0.10.2 [unknown@Linux 6.5.0-1018-azure:R 4.3.3/:memory:]
-   tissue                             n
-   &lt;chr&gt;                          &lt;dbl&gt;
- 1 blood                             17
- 2 kidney                             8
- 3 kidney blood vessel                4
- 4 lung parenchyma                    2
- 5 respiratory airway                 6
- 6 dorsolateral prefrontal cortex     1
- 7 heart left ventricle               7
- 8 bone marrow                        4
- 9 lung                               4
-10 sigmoid colon                      1
+   tissue                      n
+   &lt;chr&gt;                   &lt;dbl&gt;
+ 1 pleural effusion            1
+ 2 caudate lobe of liver       1
+ 3 cortex of kidney            7
+ 4 nose                        1
+ 5 thymus                      4
+ 6 epithelium of esophagus     1
+ 7 transverse colon            1
+ 8 liver                       1
+ 9 renal medulla               6
+10 caecum                      1
 # ℹ more rows</code></pre>
 </div>
 </div>
@@ -534,18 +534,18 @@ <h3 class="code-label">OUTPUT<i aria-hidden="true" data-feather="chevron-left"><
 </h3>
 <pre class="output" tabindex="0"><code># Source:   SQL [?? x 2]
 # Database: DuckDB v0.10.2 [unknown@Linux 6.5.0-1018-azure:R 4.3.3/:memory:]
-   assay                              n
-   &lt;chr&gt;                          &lt;dbl&gt;
- 1 sci-RNA-seq                        1
- 2 10x 5' v2                          2
- 3 10x 3' v1                          1
- 4 Smart-seq2                         1
- 5 10x 3' v3                         21
- 6 Slide-seq                          4
- 7 10x 3' v2                         27
- 8 Visium Spatial Gene Expression     7
- 9 10x 5' v1                          7
-10 scRNA-seq                          4
+   assay           n
+   &lt;chr&gt;       &lt;dbl&gt;
+ 1 10x 3' v3      21
+ 2 Slide-seq       4
+ 3 scRNA-seq       4
+ 4 Seq-Well        2
+ 5 Drop-seq        1
+ 6 10x 5' v2       2
+ 7 10x 3' v1       1
+ 8 Smart-seq2      1
+ 9 sci-RNA-seq     1
+10 10x 3' v2      27
 # ℹ more rows</code></pre>
 </div>
 </div>
@@ -740,7 +740,7 @@ <h3 class="callout-title">Exercise 1<a class="anchor" aria-label="anchor" href="
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 </button>
-<div id="collapseSolution1" class="accordion-collapse collapse" aria-labelledby="headingSolution1" data-bs-parent="#accordionSolution1">
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diff --git a/instructor/index.html b/instructor/index.html
index 6ba8907..38565e1 100644
--- a/instructor/index.html
+++ b/instructor/index.html
@@ -299,19 +299,30 @@ <h2 class="accordion-header" id="flush-headingTwelve">
  3. <a href="cell_type_annotation.html">Cell type annotation</a>
 </td>
 <td>
-TODO
+How is clustering used in scRNA-seq data analysis?<br>How are marker
+genes used to interpret clustering results?<br>What are the challenges
+and common approaches for assigning cell type labels in scRNA-seq
+data?<br>How can we further validate and refine cell type annotations
+from scRNA-seq data?
 </td>
 </tr><tr><td>
-<span class="visually-hidden">Duration: </span>01h 27m
+<span class="visually-hidden">Duration: </span>02h 00m
 </td>
 <td>
  4. <a href="multi-sample.html">Multi-sample analyses</a>
 </td>
 <td>
-TODO
+How can we identify and remove unwanted variation introduced by
+processing differences between samples (batch effects) in scRNA-seq
+data?<br>How can we identify genes that are expressed differently
+between groups of cells in scRNA-seq data (e.g., different cell types or
+conditions)?<br>How can we identify cell types (clusters) that show
+significant changes in abundance between conditions in scRNA-seq data?
+How can we address potential biases caused by differences in cell
+numbers per type?
 </td>
 </tr><tr><td>
-<span class="visually-hidden">Duration: </span>01h 39m
+<span class="visually-hidden">Duration: </span>02h 45m
 </td>
 <td>
  5. <a href="large_data.html">Working with large data</a>
@@ -322,7 +333,7 @@ <h2 class="accordion-header" id="flush-headingTwelve">
 datasets?<br>How to convert between popular single-cell data formats?
 </td>
 </tr><tr><td>
-<span class="visually-hidden">Duration: </span>01h 51m
+<span class="visually-hidden">Duration: </span>02h 57m
 </td>
 <td>
  6. <a href="hca.html">Accessing data from the Human Cell Atlas
@@ -332,7 +343,7 @@ <h2 class="accordion-header" id="flush-headingTwelve">
 How to obtain single-cell reference maps from the Human Cell Atlas?
 </td>
 </tr><tr><td>
-<span class="visually-hidden">Duration: </span>02h 21m
+<span class="visually-hidden">Duration: </span>03h 27m
 </td>
 <td>
 Finish
diff --git a/instructor/intro-sce.html b/instructor/intro-sce.html
index 0374cb5..d76a7c8 100644
--- a/instructor/intro-sce.html
+++ b/instructor/intro-sce.html
@@ -732,7 +732,7 @@ <h3 class="callout-title">Exercise 1<a class="anchor" aria-label="anchor" href="
 <button class="accordion-button hint-button collapsed" type="button" data-bs-toggle="collapse" data-bs-target="#collapseHint1" aria-expanded="false" aria-controls="collapseHint1">
   <h4 class="accordion-header" id="headingHint1">Give me a hint</h4>
 </button>
-<div id="collapseHint1" class="accordion-collapse collapse" data-bs-parent="#accordionHint1" aria-labelledby="headingHint1">
+<div id="collapseHint1" class="accordion-collapse collapse" aria-labelledby="headingHint1" data-bs-parent="#accordionHint1">
 <div class="accordion-body">
 <p>The <code>SingleCellExperiment</code> function can be used to create
 a new SingleCellExperiment object</p>
diff --git a/instructor/large_data.html b/instructor/large_data.html
index 533522c..286c130 100644
--- a/instructor/large_data.html
+++ b/instructor/large_data.html
@@ -92,11 +92,11 @@
   Orchestrating Large-Scale Single-Cell Analysis with Bioconductor
 </div>
 
-<aside class="col-md-12 lesson-progress"><div style="width: 70%" class="percentage">
-    70%
+<aside class="col-md-12 lesson-progress"><div style="width: 80%" class="percentage">
+    80%
   </div>
   <div class="progress incubator">
-    <div class="progress-bar incubator" role="progressbar" style="width: 70%" aria-valuenow="70" aria-label="Lesson Progress" aria-valuemin="0" aria-valuemax="100">
+    <div class="progress-bar incubator" role="progressbar" style="width: 80%" aria-valuenow="80" aria-label="Lesson Progress" aria-valuemin="0" aria-valuemax="100">
     </div>
   </div>
 </aside><div class="container">
@@ -707,21 +707,22 @@ <h3 class="code-label">R<i aria-hidden="true" data-feather="chevron-left"></i><i
 <h3 class="code-label">OUTPUT<i aria-hidden="true" data-feather="chevron-left"></i><i aria-hidden="true" data-feather="chevron-right"></i>
 </h3>
 <pre class="output" tabindex="0"><code>     approx
-exact   1   2   3   4   5   6   7   8   9  10  11  12  13  14
-   1   88   0   0   0   2   0   0   0   2   0   0   0   0   0
-   2    0 143   0   1   0   0   0   0   0   0   0   0   0   0
-   3    0   0  75   0   2   0   0   0   0   0   0   0   0   0
-   4    0   0   0 397   0   0   0   0   0   0   0   0   0   0
-   5    0   0   0   0 391   0   0   0   0   1   0   0   0   0
-   6    0   0   0   0   0 202   0   0   0   0   0   0   0   0
-   7    0   0   0   0   0   8 245   0   0   1   0   0   0   0
-   8    0   0   0   0   1   0   0  95   0   0   0   0   0   0
-   9    1   0   0   0   1   0   0   0 106   0   0   0   0   0
-   10   0   0   0   0   0   0   0   0   0 113   0   0   0   0
-   11   0   0   0   0   0   0   0   0   0   0 153   0   0   0
-   12   0   0   0   0   4   0   0   0   0  10   0 203   0   0
-   13   0   0   0   0   0   0   0   0   0   0   0   0 146   0
-   14   0   0   0   0   0   0   0   0   0   0   0   0   0  20</code></pre>
+exact   1   2   3   4   5   6   7   8   9  10  11  12  13  14  15
+   1   90   0   0   0   0   0   0   0   1   0   0   0   0   0   0
+   2    0 143   0   0   0   0   0   0   0   0   0   0   0   0   1
+   3    0   0  75   0   0   0   0   0   0   0   0   0   0   0   0
+   4    0   0   0 341   0   0   0   0   0   0   0   0   0   0   0
+   5    0   0   2   0 391   0   0   2   0   0   0   5   0   0   0
+   6    0   0   0   0   0  80 245   0   0   1   0   2   0   0   0
+   7    0   0   0   0   0 129   0   0   0   0   2   0   0   0   0
+   8    0   0   0   0   1   0   0  93   0   0   0   0   0   0   0
+   9    1   0   0   0   1   0   0   0 106   0   0   0   0   0   0
+   10   0   0   0   0   0   0   0   0   0 108   8   5   0   0   0
+   11   0   0   0   0   0   0   0   0   0   0 138   0   0   0   0
+   12   0   0   0   0   5   0   0   0   0   0   0 207   0   0   0
+   13   0   0   0   0   0   0   0   0   0   0   6   0 146   0   0
+   14   0   0   0   0   0   0   0   0   0   0   0   0   0  20   0
+   15   0   0   0   0   0   0   0   0   0   0   0   0   0   0  56</code></pre>
 </div>
 <p>Note that Annoy writes the NN index to disk prior to performing the
 search. Thus, it may not actually be faster than the default exact
diff --git a/instructor/multi-sample.html b/instructor/multi-sample.html
index 642eb36..73fbc82 100644
--- a/instructor/multi-sample.html
+++ b/instructor/multi-sample.html
@@ -92,11 +92,11 @@
   Orchestrating Large-Scale Single-Cell Analysis with Bioconductor
 </div>
 
-<aside class="col-md-12 lesson-progress"><div style="width: 62%" class="percentage">
-    62%
+<aside class="col-md-12 lesson-progress"><div style="width: 58%" class="percentage">
+    58%
   </div>
   <div class="progress incubator">
-    <div class="progress-bar incubator" role="progressbar" style="width: 62%" aria-valuenow="62" aria-label="Lesson Progress" aria-valuemin="0" aria-valuemax="100">
+    <div class="progress-bar incubator" role="progressbar" style="width: 58%" aria-valuenow="58" aria-label="Lesson Progress" aria-valuemin="0" aria-valuemax="100">
     </div>
   </div>
 </aside><div class="container">
@@ -272,7 +272,7 @@ <h1>Multi-sample analyses</h1>
         
         
         
-        <p>Estimated time: <i aria-hidden="true" data-feather="clock"></i> 12 minutes</p>
+        <p>Estimated time: <i aria-hidden="true" data-feather="clock"></i> 45 minutes</p>
         
         <div class="text-end">
           <button role="button" aria-pressed="false" tabindex="0" id="expand-code" class="pull-right" data-expand="Expand All Solutions " data-collapse="Collapse All Solutions "> Expand All Solutions <i aria-hidden="true" data-feather="plus"></i></button>
@@ -287,7 +287,16 @@ <h2 class="card-header">Overview</h2>
 <div class="card-body">
 <div class="inner">
 <h3 class="card-title">Questions</h3>
-<ul><li>TODO</li>
+<ul><li>How can we identify and remove unwanted variation introduced by
+processing differences between samples (batch effects) in scRNA-seq
+data?</li>
+<li>How can we identify genes that are expressed differently between
+groups of cells in scRNA-seq data (e.g., different cell types or
+conditions)?</li>
+<li>How can we identify cell types (clusters) that show significant
+changes in abundance between conditions in scRNA-seq data? How can we
+address potential biases caused by differences in cell numbers per
+type?</li>
 </ul></div>
 </div>
 </div>
@@ -295,7 +304,16 @@ <h3 class="card-title">Questions</h3>
 <div class="card-body">
 <div class="inner bordered">
 <h3 class="card-title">Objectives</h3>
-<ul><li>TODO</li>
+<ul><li>Understand the concept of batch effects and their impact on data
+analysis. Learn how to utilize tools like correctExperiment from the
+batchelor package to correct for batch effects based on sample
+information.</li>
+<li>Master the workflow for differential expression analysis (DEA),
+including grouping cells, creating pseudo-bulk samples, normalization,
+and differential expression testing with tools like edgeR.</li>
+<li>Understand differential abundance (DA) analysis, considering
+compositional effects and strategies to address them, for robust
+identification of cell type abundance changes between conditions.</li>
 </ul></div>
 </div>
 </div>
diff --git a/intro-sce.html b/intro-sce.html
index c3683c4..71a2473 100644
--- a/intro-sce.html
+++ b/intro-sce.html
@@ -730,7 +730,7 @@ <h3 class="callout-title">Exercise 1<a class="anchor" aria-label="anchor" href="
 <button class="accordion-button hint-button collapsed" type="button" data-bs-toggle="collapse" data-bs-target="#collapseHint1" aria-expanded="false" aria-controls="collapseHint1">
   <h4 class="accordion-header" id="headingHint1">Give me a hint</h4>
 </button>
-<div id="collapseHint1" class="accordion-collapse collapse" data-bs-parent="#accordionHint1" aria-labelledby="headingHint1">
+<div id="collapseHint1" class="accordion-collapse collapse" aria-labelledby="headingHint1" data-bs-parent="#accordionHint1">
 <div class="accordion-body">
 <p>The <code>SingleCellExperiment</code> function can be used to create
 a new SingleCellExperiment object</p>
diff --git a/large_data.html b/large_data.html
index 12d87f7..8a45818 100644
--- a/large_data.html
+++ b/large_data.html
@@ -92,11 +92,11 @@
   Orchestrating Large-Scale Single-Cell Analysis with Bioconductor
 </div>
 
-<aside class="col-md-12 lesson-progress"><div style="width: 70%" class="percentage">
-    70%
+<aside class="col-md-12 lesson-progress"><div style="width: 80%" class="percentage">
+    80%
   </div>
   <div class="progress incubator">
-    <div class="progress-bar incubator" role="progressbar" style="width: 70%" aria-valuenow="70" aria-label="Lesson Progress" aria-valuemin="0" aria-valuemax="100">
+    <div class="progress-bar incubator" role="progressbar" style="width: 80%" aria-valuenow="80" aria-label="Lesson Progress" aria-valuemin="0" aria-valuemax="100">
     </div>
   </div>
 </aside><div class="container">
@@ -705,21 +705,22 @@ <h3 class="code-label">R<i aria-hidden="true" data-feather="chevron-left"></i><i
 <h3 class="code-label">OUTPUT<i aria-hidden="true" data-feather="chevron-left"></i><i aria-hidden="true" data-feather="chevron-right"></i>
 </h3>
 <pre class="output" tabindex="0"><code>     approx
-exact   1   2   3   4   5   6   7   8   9  10  11  12  13  14
-   1   88   0   0   0   2   0   0   0   2   0   0   0   0   0
-   2    0 143   0   1   0   0   0   0   0   0   0   0   0   0
-   3    0   0  75   0   2   0   0   0   0   0   0   0   0   0
-   4    0   0   0 397   0   0   0   0   0   0   0   0   0   0
-   5    0   0   0   0 391   0   0   0   0   1   0   0   0   0
-   6    0   0   0   0   0 202   0   0   0   0   0   0   0   0
-   7    0   0   0   0   0   8 245   0   0   1   0   0   0   0
-   8    0   0   0   0   1   0   0  95   0   0   0   0   0   0
-   9    1   0   0   0   1   0   0   0 106   0   0   0   0   0
-   10   0   0   0   0   0   0   0   0   0 113   0   0   0   0
-   11   0   0   0   0   0   0   0   0   0   0 153   0   0   0
-   12   0   0   0   0   4   0   0   0   0  10   0 203   0   0
-   13   0   0   0   0   0   0   0   0   0   0   0   0 146   0
-   14   0   0   0   0   0   0   0   0   0   0   0   0   0  20</code></pre>
+exact   1   2   3   4   5   6   7   8   9  10  11  12  13  14  15
+   1   90   0   0   0   0   0   0   0   1   0   0   0   0   0   0
+   2    0 143   0   0   0   0   0   0   0   0   0   0   0   0   1
+   3    0   0  75   0   0   0   0   0   0   0   0   0   0   0   0
+   4    0   0   0 341   0   0   0   0   0   0   0   0   0   0   0
+   5    0   0   2   0 391   0   0   2   0   0   0   5   0   0   0
+   6    0   0   0   0   0  80 245   0   0   1   0   2   0   0   0
+   7    0   0   0   0   0 129   0   0   0   0   2   0   0   0   0
+   8    0   0   0   0   1   0   0  93   0   0   0   0   0   0   0
+   9    1   0   0   0   1   0   0   0 106   0   0   0   0   0   0
+   10   0   0   0   0   0   0   0   0   0 108   8   5   0   0   0
+   11   0   0   0   0   0   0   0   0   0   0 138   0   0   0   0
+   12   0   0   0   0   5   0   0   0   0   0   0 207   0   0   0
+   13   0   0   0   0   0   0   0   0   0   0   6   0 146   0   0
+   14   0   0   0   0   0   0   0   0   0   0   0   0   0  20   0
+   15   0   0   0   0   0   0   0   0   0   0   0   0   0   0  56</code></pre>
 </div>
 <p>Note that Annoy writes the NN index to disk prior to performing the
 search. Thus, it may not actually be faster than the default exact
diff --git a/md5sum.txt b/md5sum.txt
index a7aba31..51f66df 100644
--- a/md5sum.txt
+++ b/md5sum.txt
@@ -6,8 +6,8 @@
 "links.md" "8184cf4149eafbf03ce8da8ff0778c14" "site/built/links.md" "2024-05-02"
 "episodes/intro-sce.Rmd" "7bec0979f3e4076bb2033a132746db43" "site/built/intro-sce.md" "2024-05-02"
 "episodes/eda_qc.Rmd" "e7f8d8316c5b185864120a53f4913ece" "site/built/eda_qc.md" "2024-05-02"
-"episodes/cell_type_annotation.Rmd" "8853c2680ab7e343e249f98ad4adfe26" "site/built/cell_type_annotation.md" "2024-05-02"
-"episodes/multi-sample.Rmd" "c95671a8765365f9802da9aedadf0e3a" "site/built/multi-sample.md" "2024-05-02"
+"episodes/cell_type_annotation.Rmd" "26dd249dcbc49908b69f51857d0af997" "site/built/cell_type_annotation.md" "2024-05-02"
+"episodes/multi-sample.Rmd" "8e8baaf84d3afc32d77e5177658b1a5e" "site/built/multi-sample.md" "2024-05-02"
 "episodes/large_data.Rmd" "2735a12a1130dea13a98f4618f560ecc" "site/built/large_data.md" "2024-05-02"
 "episodes/hca.Rmd" "77d772ef05ed56e1cf5e8ffa147019ac" "site/built/hca.md" "2024-05-02"
 "instructors/instructor-notes.md" "cae72b6712578d74a49fea7513099f8c" "site/built/instructor-notes.md" "2024-05-02"
diff --git a/multi-sample.html b/multi-sample.html
index 46f976f..e027ec3 100644
--- a/multi-sample.html
+++ b/multi-sample.html
@@ -92,11 +92,11 @@
   Orchestrating Large-Scale Single-Cell Analysis with Bioconductor
 </div>
 
-<aside class="col-md-12 lesson-progress"><div style="width: 62%" class="percentage">
-    62%
+<aside class="col-md-12 lesson-progress"><div style="width: 58%" class="percentage">
+    58%
   </div>
   <div class="progress incubator">
-    <div class="progress-bar incubator" role="progressbar" style="width: 62%" aria-valuenow="62" aria-label="Lesson Progress" aria-valuemin="0" aria-valuemax="100">
+    <div class="progress-bar incubator" role="progressbar" style="width: 58%" aria-valuenow="58" aria-label="Lesson Progress" aria-valuemin="0" aria-valuemax="100">
     </div>
   </div>
 </aside><div class="container">
@@ -285,7 +285,16 @@ <h2 class="card-header">Overview</h2>
 <div class="card-body">
 <div class="inner">
 <h3 class="card-title">Questions</h3>
-<ul><li>TODO</li>
+<ul><li>How can we identify and remove unwanted variation introduced by
+processing differences between samples (batch effects) in scRNA-seq
+data?</li>
+<li>How can we identify genes that are expressed differently between
+groups of cells in scRNA-seq data (e.g., different cell types or
+conditions)?</li>
+<li>How can we identify cell types (clusters) that show significant
+changes in abundance between conditions in scRNA-seq data? How can we
+address potential biases caused by differences in cell numbers per
+type?</li>
 </ul></div>
 </div>
 </div>
@@ -293,7 +302,16 @@ <h3 class="card-title">Questions</h3>
 <div class="card-body">
 <div class="inner bordered">
 <h3 class="card-title">Objectives</h3>
-<ul><li>TODO</li>
+<ul><li>Understand the concept of batch effects and their impact on data
+analysis. Learn how to utilize tools like correctExperiment from the
+batchelor package to correct for batch effects based on sample
+information.</li>
+<li>Master the workflow for differential expression analysis (DEA),
+including grouping cells, creating pseudo-bulk samples, normalization,
+and differential expression testing with tools like edgeR.</li>
+<li>Understand differential abundance (DA) analysis, considering
+compositional effects and strategies to address them, for robust
+identification of cell type abundance changes between conditions.</li>
 </ul></div>
 </div>
 </div>
diff --git a/pkgdown.yml b/pkgdown.yml
index 648a5a3..f501a67 100644
--- a/pkgdown.yml
+++ b/pkgdown.yml
@@ -2,5 +2,5 @@ pandoc: 3.1.11
 pkgdown: 2.0.9
 pkgdown_sha: ~
 articles: {}
-last_built: 2024-05-02T17:55Z
+last_built: 2024-05-02T18:15Z