Anti-CD16 antibody [EPR22409-124]

• IHC-P

Unknown

Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-CD16 antibody [EPR22409-124] (AB246222)

Immunohistochemical analysis of paraffin-embedded human spleen tissue labeling CD16 with ab252905 at 1/500 dilution, followed by Rabbit specific IHC polymer detection kit HRP/DAB (ab209101). Positive staining on red pulp of human spleen (PMID : 26512139; 29692344). Counter stained with hematoxylin.

Secondary antibody only control : Used PBS instead of primary antibody, secondary antibody is Rabbit specific IHC polymer detection kit HRP/DAB (ab209101).

Heat mediated antigen retrieval with Tris-EDTA buffer (pH 9.0, epitope retrieval solution 2) for 20 minutes.

The section was incubated with ab252905 for 10 minutes at 37°C. The immunostaining staining was performed on a Leica Biosystems BOND^® RX instrument.

This data was developed using the same antibody clone in a different buffer formulation containing PBS, BSA, glycerol, and sodium azide (ab187039).

• ICC/IF

Unknown

Immunocytochemistry/ Immunofluorescence - Anti-CD16 antibody [EPR22409-124] (AB246222)

Immunofluorescent analysis of 4% paraformaldehyde-fixed, 0.1% Triton X-100 permeabilized human PBMCs (human peripheral blood mononuclear cells) or Raji (human Burkitt's lymphoma cell line) cell line labeling CD16 with ab246222 at 1/100 dilution, followed by Goat Anti-Rabbit IgG H&L (Alexa Fluor^® 488) (ab150077) secondary antibody at 1/1000 dilution (green). Confocal image showing membranous staining in subsets of human PBMC cells is observed. The nuclear counter stain is DAPI (blue). Tubulin is detected with Anti-alpha Tubulin antibody [DM1A] - Microtubule Marker (Alexa Fluor^® 594) (ab195889) at 1/200 dilution (red).

Secondary antibody only control : Used PBS instead of primary antibody, followed by Goat Anti-Rabbit IgG H&L (Alexa Fluor^® 488) (ab150077) secondary antibody at 1/1000 dilution.

Negative control : Raji (PMID : 11207281).

• Flow Cyt

Supplier Data

Flow Cytometry - Anti-CD16 antibody [EPR22409-124] (AB246222)

Negative control : Raji. (PMID : 11207281).

Raji (human Burkitt's lymphoma cell line) were stained with Rabbit IgG, monoclonal [EPR25A] - Isotype Control (ab172730)(Left) or ab252905 (Right) followed by a Goat anti rabbit IgG (Dylight ^® 488) at 1/2000 dilution.

Gated on viable cells.

• Flow Cyt

Unknown

Flow Cytometry - Anti-CD16 antibody [EPR22409-124] (AB246222)

Human PBMCs (human peripheral blood mononuclear cells) were stained with Rabbit IgG, monoclonal [EPR25A] - Isotype Control (ab172730) (Left) or ab246222 (Right) followed by a Goat anti rabbit IgG (Dylight ^® 488) at 1/2000 dilution. They were then stained with Alexa Fluor^® 647-conjugated anti-CD14. The expression pattern is consistent with that described in the literature. (PMID : 21738687)

Gated on viable cells.

• IP

Unknown

Immunoprecipitation - Anti-CD16 antibody [EPR22409-124] (AB246222)

CD16 was immunoprecipitated from 0.35 mg of human spleen lysate with ab252905 at 1/30 dilution. Western blot was performed from the immunoprecipitate using ab252905 at 1/1000 dilution. VeriBlot for IP Detection Reagent (HRP) (ab131366), was used as secondary antibody at 1/5000 dilution.

Lane 1 :  Human spleen lysate 10 μg (Input).
Lane 2 : ab252905 IP in human spleen lysate.
Lane 3 :  Rabbit monoclonal IgG (ab172730) instead of ab252905 in human spleen lysate.

Blocking and dilution buffer and concentration : 5% NFDM/TBST.
Exposure time : 5 seconds.

This data was developed using the same antibody clone in a different buffer formulation containing PBS, BSA, glycerol, and sodium azide (ab246222).

All lanes:

Immunoprecipitation - Anti-CD16 antibody [EPR22409-124] (ab246222)

Predicted band size: 29 kDa

Observed band size: 50-70 kDa

false

• WB

Unknown

Western blot - Anti-CD16 antibody [EPR22409-124] (AB246222)

Blocking and dilution buffer : 5% NFDM/TBST.

Exposure times.

Lane 1 : 92 seconds.

Lanes 2 & 3 : 3 minutes.

The expression profile/ molecular weight observed is consistent with what has been described in the literature (PMID : 2526846).

Negative control : Raji (PMID : 11207281).

All lanes:

Western blot - Anti-CD16 antibody [EPR22409-124] (ab246222) at 1/1000 dilution

Lane 1:

Human spleen lysate at 20 µg

Lane 2:

Human colon cancer lysate at 20 µg

Lane 3:

Raji (human Burkitt's lymphoma cell line) whole cell lysate at 20 µg

Secondary

All lanes:

Western blot - VeriBlot for IP Detection Reagent (HRP) (<a href='/en-us/products/reagents/veriblot-for-ip-detection-reagent-hrp-ab131366'>ab131366</a>) at 1/100000 dilution

Predicted band size: 29 kDa

Observed band size: 50-70 kDa

false