Application
Immunocytochemistry/ Immunofluorescence
Sample
Rat Cell (RAW267.4 macrophage)
Specification
RAW267.4 macrophage
Fixative
Paraformaldehyde
Blocking step
Serum as blocking agent for 20 minute(s) · Concentration: 10% · Temperature: 24°C
Other product details
Dilution
1/1000
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 AF555
Host species: Donkey
Clonality: Polyclonal
Conjugation: Alexa Fluor® 555
Host species: Donkey
Clonality: Polyclonal
Conjugation: Alexa Fluor® 555
Dilution
1/1000
Additional data
Additional Notes
Macrophage cell line RAW264.7 was grown in DMEM/10% FCS and grown on glass cover slips in 6 well plates.
Fixed in 1%PFA prior to staining.
Wash 3x3min in PBS.
Block 10% donkey serum for 20min at RT.
Primary Ab diluted 1/1000 in PBS and incubated overnight at 4deg, also works by incubating for 2 hours at RT.
Wash 3x3min in PBS.
Secondary - donkey anti-rat AF555 (yellow) 1/1000 in PBS for 1h at RT.
Wash 3x3min in PBS
Counterstained with Hoechst 33258 (blue) for 10mins at RT.
Wash 3min in dH2O
Coverslips removed tapped dry and placed on glass slides and mounted in Invitrogen ProLong gold and kept at 4deg until imaged.
Fixed in 1%PFA prior to staining.
Wash 3x3min in PBS.
Block 10% donkey serum for 20min at RT.
Primary Ab diluted 1/1000 in PBS and incubated overnight at 4deg, also works by incubating for 2 hours at RT.
Wash 3x3min in PBS.
Secondary - donkey anti-rat AF555 (yellow) 1/1000 in PBS for 1h at RT.
Wash 3x3min in PBS
Counterstained with Hoechst 33258 (blue) for 10mins at RT.
Wash 3min in dH2O
Coverslips removed tapped dry and placed on glass slides and mounted in Invitrogen ProLong gold and kept at 4deg until imaged.
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提交于 Jun 07 2012