FITC staining of polar bodies?

Model Organism: Worms Scientific Discussion
1

last week I did some FITC staining of different stage eggs and noticed that in addition to the normal outline of the egg shell, prominent bright spots on the surface of different stage eggs.

Some had two, some one and at least one had three. It can’t be an artefact as the positioning seems relatively conserved, so I was wondering whether they were stained polar bodies…but then shouldn’t the egg be effectively impermeable to FITC in the later stages?

At least one of the eggs show at the links below is late stage.

http://imgur.com/o0IUx
http://imgur.com/IwIL7
http://imgur.com/0nuA7
http://imgur.com/RVDij

Anyone cast light on what is probably a frequent observation to trained embryologists.

P.S. apologies for the poor quality of the images, we have no DIC and the inverted scope is limited in it’s capacity to resolve bright field. We’re saving up our pennies for a good scope.

Steve

2

My thanks to Bob Goldstein (pers. comm.) for confirming that I was most likely seeing FITC staining of the 1st (outer) polar body, although he thought the 2nd would not normally be stained (I’ll check whether my observations of two spots are replicable).

Bob also pointed me to a paper out of the Oegema lab;

http://www.ncbi.nlm.nih.gov/pubmed/22908315

one of their observations perhaps explains why most people don’t see this phenomenon. Apparently bleaching as opposed to cut 'n release effectively makes the eggs impermeable to staining (the polar bodies), so mass screening for egg shell mutants probably wouldn’t pick this up either…

Steve