when tumor bx slides are sent for immunohistochemistry,after few weeks it starts to fade out so slides cant be. read then.Is there any option to prevent fading?
We are adopting manual way of IHC staining.First antigen retrieval,then pry antibody followed by sec antibody.In between buffer washes are given.
Hello
You can use Fluoromount G, which keeps fluorescence for a really long time. Also, you can mount in PBS & keep them at- 20°C (darkness).
Best
It sounds like you don't do your immunohistochemistry yourself but rather receive slides stained elsewhere. Without knowing any deatils of how your slides are stained, one thing you can do, is store your slides in the dark.
IHC slides don't fade out, because is a chemical staining. I think that you talking about IF, if yes, you need to used one anti-fadding reagent before mount your slide. I usually use prolong and the signal remains at least for a month, if you stock you slide in the dark.
If you mean IF, it will fade after one wk unless you put slides in dark light and at -20 C
If you mean IHC for paraffin slide, it will be ok for more than 1 year specially if you use permanent mountant.
As several others have written first step is to store the slides in the dark and cool. 4 deg C is best.
Next step since you are not staining and I assume mounting the coverslips yourself, you need to get the lab or service to use an anti-fade agent in the mounting medium. This is the most critical. Presuming that the service lab will work with you, you can buy for them such an agents to use on your slides. There are two commercial sources - Vector Labs has a product which is quite economical - Vectashield. They market several levels of anti-fade agents depending on how long you want to store the slides for. Another mounting medium is produced by LifeTechnologies (formerly Molecular Probes) - Prolong. Prolong also comes in several levels of protection. Prolong has the advantage that is hardens to permanent mounting media.
Vectashield can be purchased in a hardening form or not. An advantage to the non-hardening formulas is that you can unmount the coverslip and retrieve the section or specimen for DNA extraction or whatsoever. The hardening mounting medias preserve material for years. And also make shipping completed slides easy.
You should use prolong gold anti fade reagent to mount cover slip, keep your slides in dark and -20C, and also try to use cy3 (red), instead of alexa 488 (green), cy3 (red) is more stable as comare to Alexa 488 (green), I hope it will work for you.
1. Use of anti-fade reagents before mounting. 2. Storing the slides in a light-tight container at 4 C. I think storing them at -20 can lead to dehydration of all the solutions used in the process and giving background staining. As mentioned by other researchers, fading takes place mostly in IF staining and not so much in IHC staining. Hope it helps.
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