Application
Immunocytochemistry/ Immunofluorescence
Sample
Mouse Cell (Spleen)
Specification
Spleen
Fixative
Paraformaldehyde
Blocking step
Serum as blocking agent for 30 minute(s) · Concentration: 10% · Temperature: 24°C
Other product details
Dilution
1/10
Incubation time
16 hour(s) and 0 minute(s) · Temperature: 4°C · Diluent: PBS
Secondary antibody
Name
Non-Abcam antibody was used: Invitrogen Donkey anti-Rat AF488
Host species: Donkey
Clonality: Polyclonal
Conjugation: Alexa Fluor® 488
Host species: Donkey
Clonality: Polyclonal
Conjugation: Alexa Fluor® 488
Dilution
1/1000
Additional data
Additional Notes
7um non-perfused rat spleen, post-fixed
Fix - 4%PFA 10mins @ RT
Wash - 3x3min PBS
Block - 5% Normal Donkey Serum in PBS - 30mins @ RT
Tap-off Block - except for negative
Primary F4/80 1/100 in PBS - works well at 2 hours @ RT or 4deg overnight (image shown)
Wash - 3x3min PBS
Secondary - Donkey Anti-Rat AF488 (Green) 1/1000 in PBS for 1 hour @ RT
Wash - 3x3min PBS
Counterstain - Hoescht 33342 (Blue)
Final Wash - 1x3min dH2O
Mount - Pro-long gold
Kept at 4deg prior to imaging
Negative control - without primary antibody
Fix - 4%PFA 10mins @ RT
Wash - 3x3min PBS
Block - 5% Normal Donkey Serum in PBS - 30mins @ RT
Tap-off Block - except for negative
Primary F4/80 1/100 in PBS - works well at 2 hours @ RT or 4deg overnight (image shown)
Wash - 3x3min PBS
Secondary - Donkey Anti-Rat AF488 (Green) 1/1000 in PBS for 1 hour @ RT
Wash - 3x3min PBS
Counterstain - Hoescht 33342 (Blue)
Final Wash - 1x3min dH2O
Mount - Pro-long gold
Kept at 4deg prior to imaging
Negative control - without primary antibody
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投稿 Aug 17 2011