Application
Immunocytochemistry
Sample
Human Cell (Kelly Neuroblastoma cell line)
Specification
Kelly Neuroblastoma cell line
Fixative
Paraformaldehyde
Permeabilization
No
Blocking step
Serum as blocking agent for 20 minute(s) · Concentration: 10% · Temperature: 24°C
Other product details
Dilution
1/200
Incubation time
2 hour(s) and 0 minute(s) · Temperature: 24°C · Diluent: PBS
Secondary antibody
Name
Non-Abcam antibody was used: Vector Goat Anti Mouse Biotinylated
Host species: Goat
Clonality: Polyclonal
Conjugation: Biotin
Host species: Goat
Clonality: Polyclonal
Conjugation: Biotin
Dilution
1/200
Additional data
Additional Notes
Kelly Neuroblastoma cell line was cultured in RPMI/10% FCS on coverslips in 12 well plates.
Cells were washed with PBS and fixed in 4% PFA until staining.
Staining protocol:
Wash 2x2min PBS;
Endogenous peroxidase block - 3%H2O2 in methanol for 15mins at RT;
Wash 2x2min PBS;
Serum block - 10% Goat serum;
Primary - tap off serum block add primary in PBS for 2h at RT;
Wash 2x2min PBS;
Secondary - Goat Anti Mouse Biotin (Vector) 5ul/ml in 0.1% PBSA 30mins at RT;
Wash 2x2min PBS;
ABC from Vector 30min at RT;
Wash 2x2mins PBS;
DAB (Vector) (brown);
Wash 2x2min H2O;
Counterstain - Harris' Haematoxylin (purple);
Wash off with H2O;
Dehydrate 2mins - 1x75%EtOh 2x100%EtOh 2xHistoclear;
Mount using Histomount.
Cells were washed with PBS and fixed in 4% PFA until staining.
Staining protocol:
Wash 2x2min PBS;
Endogenous peroxidase block - 3%H2O2 in methanol for 15mins at RT;
Wash 2x2min PBS;
Serum block - 10% Goat serum;
Primary - tap off serum block add primary in PBS for 2h at RT;
Wash 2x2min PBS;
Secondary - Goat Anti Mouse Biotin (Vector) 5ul/ml in 0.1% PBSA 30mins at RT;
Wash 2x2min PBS;
ABC from Vector 30min at RT;
Wash 2x2mins PBS;
DAB (Vector) (brown);
Wash 2x2min H2O;
Counterstain - Harris' Haematoxylin (purple);
Wash off with H2O;
Dehydrate 2mins - 1x75%EtOh 2x100%EtOh 2xHistoclear;
Mount using Histomount.
The reviewer received a reward from Abcam’s Loyalty Program in thanks for submitting this Abreview and for helping the scientific community make better-informed decisions.
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投稿 Sep 28 2012