[Histonet] Liquid cover glass for H&E stained cell culture in 6 well plates; toluidine blue stain for cell cultures

[gayle callis]

What are people using as a liquid cover glass for stained cells in 6 well
culture plates to be view with inverted microscope with having that annoying
refractile, funky look to the cells.  

 

Cells are grown in 6 well culture plates and with previous cultures, been
stained with Hema 3 (hematological stain) and Giemsa of which neither has
been successful to see cell membranes and nuclei clearly.   Viewing is done
with inverted microscope and cells are counted.   

 

I know there are some stains that can be viewed left under water with
inverted scope.  H&E obviously cannot be left under water due to eosin
washing out of cytoplasm.   

 

A possibility is  letting H&E or T blue stained cells dry, then put a thin
layer of permanent mounting media to flow over the cells to fill in spaces,
let dry and view with inverted microscope.  I think one could do an old
bonehead trick we used on surface stained, thicker ground MMA embedded
undecalcified bone, flood with immersion oil and put a cover glass on top.
Very messy!!!   However one could let H&E dry after last alcohol rinse,  put
a layer of immersion oil over the cells, and view with inverted microscope.
Anything to fill in the cellular spaces on a confluent sheet of cells but
let the oil be the cover glass.  Growing cells on a cover glass or on slide
with wells/gaskets is not an option.   

 

Any suggested stains with protocols would be appreciated too with thoughts
here about using Toluidine blue.  The recipe for making up the stain,
staining method and preferred fixative would be appreciated.   You can send
information privately if you wish.  

 

A challenge to be sure

 

Thanks 

 

Gayle Callis

HTL/HT/MT(ASCP)