Recombinant Anti-Choline Acetyltransferase antibody [EPR13024(B)] (ab181023)

This antibody recognizes all Choline Acetyltransferase isoforms with the MW ranging from 70-82 KDa based on immunogen blast.

Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) analysis of Human cerebrum tissue sections labeling Choline Acetyltransferase with purified ab181023 at 1/2000 dilution (0.28 µg/ml). Heat mediated antigen retrieval was performed using heat mediated antigen retrieval using ab93684 (Tris/EDTA buffer, pH 9.0). ImmunoHistoProbe one step HRP Polymer (ready to use) was used as the secondary antibody. Negative control: PBS instead of the primary antibody. Hematoxylin was used as a counterstain.

Immunocytochemistry/ Immunofluorescence analysis of U-87 MG (Human glioblastoma-astrocytoma epithelial cell) cells labeling Choline Acetyltransferase with purified ab181023 at 1/50 dilution (10 µg/ml). Cells were fixed in 4% Paraformaldehyde and permeabilized with 0.1% tritonX-100. Cells were counterstained with ab195889 Anti-alpha Tubulin antibody [DM1A] - Microtubule Marker (Alexa Fluor^® 594) at 1:200 (2.5 µg/ml). Goat anti rabbit IgG (Alexa Fluor^® 488, ab150077) was used as the secondary antibody at 1/1000 (2 µg/ml) dilution. DAPI (blue) was used as nuclear counterstain. PBS instead of the primary antibody was used as the secondary antibody only control.

Blocking/ Dilution buffer: 5% NFDM /TBST.

Flow Cytometry analysis of SH-SY5Y (Human neuroblastoma epithelial cell) cells labeling Choline Acetyltransferase with purified ab181023 at 1/60 dilution (10 µg/ml) (red). Cells were fixed with 4% Paraformaldehyde and permeabilised with 90% Methanol. A Goat anti rabbit IgG (Alexa Fluor^® 488, ab150077) secondary antibody was used at 1/2000. Isotype control - Rabbit monoclonal IgG (Black). Unlabeled control - Cell without incubation with primary antibody and secondary antibody (Blue).

ab181023 (purified) at 1/30 dilution (2ug) immunoprecipitating Choline Acetyltransferase in Human fetal brain lysate. Human fetal brain lysate 10ug
Lane 2 (+): ab181023 & Human fetal brain lysate
Lane 3 (-): Rabbit monoclonal IgG (ab172730) instead of ab181023 in Human fetal brain lysate
For western blotting, VeriBlot for IP secondary antibody (HRP) (ab131366) was used at 1/1000 dilution.
Blocking and diluting buffer: 5% NFDM/TBST.

Immunofluorescent analysis of 4% paraformaldehyde fixed SH-SY5Y cells labeling Choline Acetyltransferase using ab181023 (unpurified) at a 1/100 dilution. A Goat anti rabbit IgG (Alexa Fluor®555) was used as the secondary at a 1/100 dilution.

Flow Cytometry analysis of SH-SY5Y cells labeling Choline Acetyltransferase using ab181023 (unpurified) at a 1/110 dilution (pink). Goat anti rabbit IgG (FITC) used as the secondary at a 1/150 dilution. Isotype control Rabbit monoclonal IgG (green). Cells were fixed in 2% paraformaldehyde.

Lysate from Human fetal brain (Lane 1) and  negative control (Lane 2) were immunoprecipitated with ab181023 (unpurified) at a 1/70 dilution. A anti-rabbit IgG (HRP), specific to the non-reduced form of IgG at a 1/1500 dilution for the secondary. Blocking/ Dilution buffer: 5% NFDM/TBST.