Brain organoid

Due to current restrictions to limit the spread of the new Coronavirus SARS-CoV-2, our collaborators at the Italian Institute of Technology (IIT)-CLNS SAPIENZA shared with us their samples, brain organoid sections of 50 μm thickness, and we made acquisitions CrestOptics with X-Light V3 Confocal Spinning Disk.

In particular, brain organoid structures shown here derive from in-house derived control iPSC line grown for 71 days under continuous shaking on a 3D orbital shaker in order to ensure the homogeneous diffusion of the media nutrients within the structures. Then, organoids were fixed, sliced and sections were stained for various differentiation markers (described in Figure A and Figure B) in order to confirm the generation of specific cortical neuron subtypes.

Figure A)

  • MAP2 (Alexa 594 fluorophore, shown in red), a microtubule-associated protein present in the cytoskeleton of neuronal dendrites.
  • CTIP2 (Alexa 488 fluorophore, shown in green), which stains cortical neuron nuclei that localize in the layer V of the cortical plate and its expression is involved in the early specification of a distinct class of cerebral projection neurons.

Stitched image

(Plan Apochromat Lambda 60X oil, 1.45 NA)

3D volume reconstruction and orthogonal (XZ, YZ) views

(Plan Apochromat Lambda 60X oil, 1.45 NA)

Figure B)

  • TBR1 (Alexa 750 fluorophore, shown in magenta) a nuclear transcription factor important for the glutamatergic neuron differentiation during the cortical development and its staining is specific for those neurons that localize in layer VI of the cortical plate.
  • MAP2 (Alexa 488 fluorophore, shown in green), a microtubule-associated protein present in the cytoskeleton of neuronal dendrites.

Stitched image

(Plan Apochromat Lambda 60X oil, 1.45 NA)

Maximum Intensity Projection and 3D volume reconstruction

(Plan Apochromat Lambda 60X oil, 1.45 NA)

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