RE: Immunostaining cells on Matrigel -- how we do it
Charlene and interested others:
Immunofluorescent staining can be accomplished on 24 well plates, if you have an inverted fluorescent or confocal microscope with which to view the stained plates. Sometimes the availability of the appropriate end product observation tool limits what we do in the lab for our investigators. Hence the proliferation of many useful product to grow and observe cultured cells. Below are a few we have used in our lab.
Chamber slides: Available from Nunc, Fisher Scientific and probably others. These slides come in single well up to 16 well formats with various options for growth surface treatment, e.g. matrigel, fibronectin, collagen type I etc. These slides have a gasket and plastic well that contains the cell media for growth. After cells have been grown on the slides, we fix with 10 % NBF and stain using standard immunofluorescent techniques. Remove the chamber wells and gasket, apply fluorescent mounting media e.g. vectashield and No 1 cover slip of sufficient size. Note: we use no 1 (thickness) cover glass for all fluorescent applications because the resultant images are sharper.
BioCoat inserts: We get ours from Fisher. These cylindrical inserts have a porous membrane on the bottom impregnated with matrigel, fibronectin, collagen type I, etc. They fit into standard multiwell tissue culture plates. Cells grown on these inserts may adhere to the membrane or pass through it to the plate below. Many investigators use these to assess the invasive capacity of tumor cells in vitro. The good news is that these inserts can be, fixed and stained in place or on another plate according the investigative question. The membrane can be cut out using a Number 11 scalpel blade or exacto knife floated onto a standard microscope slide, cover slipped, mounted and viewed on a standard scope.
Of course, if you have access to an inverted fluorescent or confocal scope, you can fix and stain your cultured cells in the plate and view them directly simplifying matters greatly.
Hope this helps, Donna Montague, Center for Orthopaedic Research, UAMS, Little Rock, AR
From: Carlene L. Zindl [mailto:firstname.lastname@example.org]
Sent: Tuesday, November 26, 2002 4:16 PM
Subject: Immunostaining cells on Matrigel
Does anyone have a protocol or good reference for performing immunofluorescent staining on cells grown on Matrigel? Should the cells be grown on coverslips or is the staining possible in 24 well plates? Thank you for any info and I hope everyone has a happy Thanksgiving!
St. Louis, MO
<< Previous Message | Next Message >>