Figure 8.

Quantification of compound mRNA puncta in DRG neurons using FISH-quant

(A) Obtain the image of “average punctum” by averaging all discrete mRNA puncta from the same batch of images. Image quality improves as more puncta are averaged. Left: Images of the ‘average puncta’ with maximum intensity projections along the three axes. Right: Images of the ‘average puncta’ subjected to Gaussian fit along the XYZ axes. Size of the averaging region in this example: XY = 9 (pixel), Z = 9 (pixel).

(B) An estimate calling of discrete puncta number quantified within a compound puncta by PSF superposition approach along the three axis with the ‘average puncta’. Using the PSF superposition approach, the compound punctum is reconstructed. The software trials placement of ‘average puncta’ until the best description of the compound puncta is obtained.