Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) analysis of human tonsil tissue sections labeling IgG2 with purified ab134050 at 1:40000 dilution (0.025 µg/ml). Heat mediated antigen retrieval was performed 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.

Flow Cytometry analysis of Raji (human Burkitt's lymphoma cell line) cells labeling IgG2 with purified ab134050 at 1:100 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).

Immunocytochemistry/ Immunofluorescence analysis of Raji (human Burkitt's lymphoma cell line) cells labeling IgG2 with purified ab134050 at 1:100 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) 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.

All lanes : Anti-IgG2 antibody [EPR4418] (ab134050) at 1/5000 dilution (Purified)

Lane 1 : Human spleen lysates
Lane 2 : Human tonsil lysates

Lysates/proteins at 15 µg per lane.

Secondary
All lanes : Goat Anti-Rabbit IgG (HRP) with minimal cross-reactivity with human IgG at 1/2000 dilution

Predicted band size: 36 kDa
Observed band size: 52 kDa why is the actual band size different from the predicted?

All lanes : Anti-IgG2 antibody [EPR4418] (ab134050) at 1/10000 dilution (Unpurified)

Lane 1 : Human Recombinant IgG2 at 0.01 µg
Lane 2 : Human Recombinant IgG1 at 0.01 µg
Lane 3 : Human Recombinant IgG3 at 0.01 µg
Lane 4 : Human Recombinant IgG4 at 0.01 µg
Lane 5 : Human spleen lysate at 10 µg
Lane 6 : Human tonsil lysate at 10 µg

Secondary
All lanes : HRP labelled goat anti-rabbit at 1/2000 dilution

Predicted band size: 36 kDa

Immunohistochemical analysis of paraffin-embedded human tonsil tissue labeling IgG2 with unpurified ab134050 at 1/500 dilution.

Perform heat mediated antigen retrieval before commencing with IHC staining protocol.

Immunohistochemical analysis of paraffin-embedded human thyroid gland tissue labeling IgG2 with unpurified ab134050 at 1/500 dilution.

Perform heat mediated antigen retrieval before commencing with IHC staining protocol.