diff --git a/workflows/imaging/fluorescence-nuclei-segmentation-and-counting/.dockstore.yml b/workflows/imaging/fluorescence-nuclei-segmentation-and-counting/.dockstore.yml
new file mode 100644
index 0000000000..e51c75ebe2
--- /dev/null
+++ b/workflows/imaging/fluorescence-nuclei-segmentation-and-counting/.dockstore.yml
@@ -0,0 +1,12 @@
+version: 1.2
+workflows:
+- name: main
+  subclass: Galaxy
+  publish: true
+  primaryDescriptorPath: /segmentation-and-counting.ga
+  testParameterFiles:
+  - /segmentation-and-counting-tests.yml
+  authors:
+  - name: Leonid Kostrykin
+    orcid: 0000-0003-1323-3762
+    url: https://github.com/kostrykin/
diff --git a/workflows/imaging/fluorescence-nuclei-segmentation-and-counting/CHANGELOG.md b/workflows/imaging/fluorescence-nuclei-segmentation-and-counting/CHANGELOG.md
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index 0000000000..2169bdc88b
--- /dev/null
+++ b/workflows/imaging/fluorescence-nuclei-segmentation-and-counting/CHANGELOG.md
@@ -0,0 +1,5 @@
+# Changelog
+
+## [0.1] - 2024-02-29
+
+- Creation of workflow for segmentation and counting of cell nuclei in fluorescence microscopy images.
diff --git a/workflows/imaging/fluorescence-nuclei-segmentation-and-counting/README.md b/workflows/imaging/fluorescence-nuclei-segmentation-and-counting/README.md
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index 0000000000..60d0a12344
--- /dev/null
+++ b/workflows/imaging/fluorescence-nuclei-segmentation-and-counting/README.md
@@ -0,0 +1,17 @@
+# Segmentation and counting of cell nuclei in fluorescence microscopy images
+
+This workflow performs segmentation and counting of cell nuclei using fluorescence microscopy images. The segmentation step is performed using Otsu thresholding (Otsu, 1979). The workflow is based on the tutorial: https://training.galaxyproject.org/training-material/topics/imaging/tutorials/imaging-introduction/tutorial.html
+
+![](test-data/overlay_image.png)
+
+## Inputs
+
+**`input_image`:** The fluorescence microscopy images to be segmented. Must be the single image channel, which contains the cell nuclei.
+
+## Outputs
+
+**`overlay_image`:** An overlay of the original image and the outlines of the segmentated objects, each also annotated with a unique number.
+
+**`objects_count`:** Table with a single column `objects` and a single row (the actual number of objects).
+
+**`label_image`:** The segmentation result (label map, which contains a unique label for each segmented object).
diff --git a/workflows/imaging/fluorescence-nuclei-segmentation-and-counting/segmentation-and-counting-tests.yml b/workflows/imaging/fluorescence-nuclei-segmentation-and-counting/segmentation-and-counting-tests.yml
new file mode 100644
index 0000000000..35facddc5d
--- /dev/null
+++ b/workflows/imaging/fluorescence-nuclei-segmentation-and-counting/segmentation-and-counting-tests.yml
@@ -0,0 +1,19 @@
+- doc: Test outline for segmentation-and-counting.ga
+  job:
+    input_image:
+      class: File
+      path: test-data/input_image.tiff
+      filetype: tiff
+  outputs:
+    overlay_image:
+      path: test-data/overlay_image.png
+      compare: sim_size
+      delta: 100
+    objects_count:
+      path: test-data/objects_count.tabular
+      compare: diff
+      lines_diff: 0
+    label_image:
+      path: test-data/label_image.tiff
+      compare: sim_size
+      delta: 100
diff --git a/workflows/imaging/fluorescence-nuclei-segmentation-and-counting/segmentation-and-counting.ga b/workflows/imaging/fluorescence-nuclei-segmentation-and-counting/segmentation-and-counting.ga
new file mode 100644
index 0000000000..962daf4de0
--- /dev/null
+++ b/workflows/imaging/fluorescence-nuclei-segmentation-and-counting/segmentation-and-counting.ga
@@ -0,0 +1,343 @@
+{
+    "a_galaxy_workflow": "true",
+    "annotation": "This workflow performs segmentation and counting of cell nuclei using fluorescence microscopy images. The segmentation step is performed using Otsu thresholding (Otsu, 1979). The workflow is based on the tutorial: https://training.galaxyproject.org/training-material/topics/imaging/tutorials/imaging-introduction/tutorial.html",
+    "creator": [
+        {
+            "class": "Person",
+            "identifier": "https://orcid.org/0000-0003-1323-3762",
+            "name": "Leonid Kostrykin",
+            "url": "https://github.com/kostrykin/"
+        }
+    ],
+    "format-version": "0.1",
+    "license": "MIT",
+    "release": "0.1",
+    "name": "Segmentation and counting of cell nuclei in fluorescence microscopy images",
+    "steps": {
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+                    "name": "input_image"
+                }
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+            "label": "input_image",
+            "name": "Input dataset",
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+}
diff --git a/workflows/imaging/fluorescence-nuclei-segmentation-and-counting/test-data/input_image.tiff b/workflows/imaging/fluorescence-nuclei-segmentation-and-counting/test-data/input_image.tiff
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diff --git a/workflows/imaging/fluorescence-nuclei-segmentation-and-counting/test-data/label_image.tiff b/workflows/imaging/fluorescence-nuclei-segmentation-and-counting/test-data/label_image.tiff
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diff --git a/workflows/imaging/fluorescence-nuclei-segmentation-and-counting/test-data/objects_count.tabular b/workflows/imaging/fluorescence-nuclei-segmentation-and-counting/test-data/objects_count.tabular
new file mode 100644
index 0000000000..1d8cebc77d
--- /dev/null
+++ b/workflows/imaging/fluorescence-nuclei-segmentation-and-counting/test-data/objects_count.tabular
@@ -0,0 +1,2 @@
+objects
+20
\ No newline at end of file
diff --git a/workflows/imaging/fluorescence-nuclei-segmentation-and-counting/test-data/overlay_image.png b/workflows/imaging/fluorescence-nuclei-segmentation-and-counting/test-data/overlay_image.png
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