Immunocytochemistry/ Immunofluorescence abreview for Anti-PCNA antibody - Proliferation Marker

Excellent
Abreviews
Application
Immunocytochemistry/ Immunofluorescence
Sample
Zebrafish Cell (Gastrula Embryo Whole Mount)
Specification
Gastrula Embryo Whole Mount
Fixative
Formaldehyde
Permeabilization
Yes - Proteinase K 10min r.t. re-fix in PFA 20min r.t. then 0.5% Triton X-100 in PBS 15min r.t.
Blocking step
2% goat serum, 2mg/mL BSA, 0.1% Tween-20 in PBS as blocking agent for 2 hour(s) and 0 minute(s) · Concentration: 2% · Temperature: 20°C

Other product details

Dilution
1/500
Incubation time
16 hour(s) and 0 minute(s) · Temperature: 4°C · Diluent: Same as Block

Secondary antibody

Name
Non-Abcam antibody was used: Goat anti-Rabbit Alexa Fluor 488
Host species: Goat
Clonality: Polyclonal
Conjugation: Alexa Fluor® 488
Dilution
1/500

Additional data

Additional Notes
Zebrafish embryos were fixed overnight at 4C when they had reached 60% epiboly. They were processed for ICC/IF as outlined above and post-fixed in PFA for 20min at r.t. after extensive washing of the secondary antibody. Pieces of lateral blastoderm were then mounted in Vectashield and image stacks through the epi- and hypoblasts were collected on a confocal microscope. The images shown are maximum intensity Z-projections of 4, 1micron slices. The embryonic cell cycle has minimal gap phases at this point, so most nuclei would be expected to be PCNA-positive. The left panel shows DAPI stained nuclei, the center panel is PCNA staining, and the right panel is the merged image.

Mr. Hank Farr

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Submitted Mar 30 2012

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