Negative staining – a spectacular job!
· One drop of suspension of 300 mesh formvar coated grid – 30 minutes
· Wick away excess suspension with paper arrow (Watman No.1)
· Place one drop of UranyLess on the grid before it is completely dry
· Stain for 3 minutes
· Wick away excess stain with paper arrow (Watman No.1)
· Allow the grid(s) to dry completely before examination on the TEM
Times can be altered (suspension time, stain time) to suit the “look” you desire.
Many thanks to EMS!
Cheers,
Debra M. Townley, Baylor College of Medicine
Thank you for you post. I m in the process of trying the Uranylless.
Loui
Hello EM Experts,
Has any one use Pre packaged Lead citrate? Are they any good. Or just the old Reynolds Lead citrate?
I will appreciate our input
Loui Harkins,
EM LAB, Univ Mississippi Medical Center
Jackson, MS
Thanks Debra for your negative stain protocol with Uranyless (non-radioactive lanthanide based). Did you do a stain intensity comparison to uranyl acetate (e.g. 2% Uac aqueous solution) and observed any decrease in negative stain intensity surrounding viral/microorganism particles? Any difference in stain precipitate between the two stains? Could you share images ?
I've tried 2.5% gadolinium triacetate and UAR-EMS for negative staining with similar results yet with slightly lower stain intensity compared to 2% Uac for AAV samples.
Some negative stain procedures have rinse steps between the sample suspension application/absorption and stain. Which could be sample type & suspension concentration related.
Philip Seifert, HTL(ASCP)
Core Manager & EM Specialist
Morphology Core
Schepens Eye Research Institute of Mass Eye and Ear
Boston, MA 02114