Generally, 4°C overnight will give more efficient and specific staining. However, many antibodies will also work very well with one or two hours incubation at room temperature. Actually, I need a short list of primary antibodies that working well at room temperature for one hour.
Most of my primary antibodies worked overnight 4°C or 1 h at room temperature. The only reason I use overnight 4°C is for saving the precious antibodies from collaborators for repeated use. You really need to try once to know whether the antibodies work in both conditions. But I never had problems in either of the incubation conditions.
Most of my primary antibodies worked overnight 4°C or 1 h at room temperature. The only reason I use overnight 4°C is for saving the precious antibodies from collaborators for repeated use. You really need to try once to know whether the antibodies work in both conditions. But I never had problems in either of the incubation conditions.
Both should work well unless the overnight incubation is not followed by efficient wash steps which will then cause a high background noise.
I recommend overnight incubation at 4°C too, and atleast 4 washes, to avoid high background. If time is limited, incubation for 1 hour, would still give you results.
depends on which marker or tissue is utilized, it is experienced that some markers in spite of manual instructions has diverse results, for example calretinin in 80 minutes has no to very weak reaction in breast tissue whereas overnight incubation is amazing.
- Badges
- Science method
- Similar topics
- Biological Science
- Histology
- Staining