Anti-CD69 antibody [EPR21814]

• IHC-P

Supplier Data

Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-CD69 antibody [EPR21814] (AB233396)

Immunohistochemical analysis of paraffin-embedded human tonsil tissue labeling CD69 with ab233396 at 1/500 dilution, followed by a ready to use Goat Anti-Rabbit IgG H&L (HRP). Membranous and cytoplasmic staining in lymphoid cells of human tonsil (PMID : 2807372) is observed. Counter stained with Hematoxylin.
Secondary antibody only control : Used PBS instead of primary antibody, secondary antibody is a ready to use Goat Anti-Rabbit IgG H&L (HRP).
Perform heat mediated antigen retrieval using ab93684 (Tris/EDTA buffer, pH 9.0).

• IHC-P

Supplier Data

Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-CD69 antibody [EPR21814] (AB233396)

Immunohistochemical analysis of paraffin-embedded human cervix cancer tissue labeling CD69 with ab233396 at 1/500 dilution, followed by a ready to use Goat Anti-Rabbit IgG H&L (HRP). Membranous and cytoplasmic staining in infiltrated lymphoid cells of human cervical cancer is observed. Counter stained with Hematoxylin.
Secondary antibody only control : Used PBS instead of primary antibody, secondary antibody is a ready to use Goat Anti-Rabbit IgG H&L (HRP).
Perform heat mediated antigen retrieval using ab93684 (Tris/EDTA buffer, pH 9.0).

• IP

Supplier Data

Immunoprecipitation - Anti-CD69 antibody [EPR21814] (AB233396)

CD69 was immunoprecipitated from 0.35 mg Daudi (human Burkitt's lymphoma lymphoblast) treated with 50 ng/ml phorbol-12-myristate-13-acetate (PMA, ab120297) for 24 hours whole cell lysate with ab233396 at 1/30 dilution. Western blot was performed from the immunoprecipitate using ab233396 at 1/5,000 dilution. VeriBlot for IP Detection Reagent (HRP) (ab131366), was used for detection at 1/5,000 dilution.

Lane 1 : Daudi (human Burkitt's lymphoma lymphoblast) treated with 50 ng/ml phorbol-12-myristate-13-acetate (PMA, ab120297) for 24 hours whole cell lysate 10 μg (Input).
Lane 2 : ab233396 IP in Daudi treated with 50 ng/ml phorbol-12-myristate-13-acetate (PMA, ab120297) for 24 hours whole cell lysate (+).
Lane 3 : Rabbit monoclonal IgG (ab172730) instead of ab233396 in Daudi treated with 50 ng/ml phorbol-12-myristate-13-acetate (PMA, ab120297) for 24 hours whole cell lysate (-).

Blocking/Dilution buffer : 5% NFDM/TBST.

All lanes:

Immunoprecipitation - Anti-CD69 antibody [EPR21814] (ab233396)

Predicted band size: 23 kDa

false

Exposure time: 30s

• WB

Lab

Western blot - Anti-CD69 antibody [EPR21814] (AB233396)

Blocking/Dilution buffer : 5% NFDM/TBST.

PMA treatment increases the basal level of p28/32 on Daudi. PMID : 1617156.

All lanes:

Western blot - Anti-CD69 antibody [EPR21814] (ab233396) at 1/5000 dilution

Lane 1:

Un-treated Daudi (human Burkitt's lymphoma lymphoblast) whole cell lysate at 10 µg

Lane 2:

Daudi treated with 50 ng/ml phorbol-12-myristate-13-acetate (PMA, <a href='/en-us/products/biochemicals/phorbol-12-myristate-13-acetate-pma-pkc-activator-ab120297'>ab120297</a>) for 24 hours at 10 µg

Secondary

All lanes:

Western blot - Goat Anti-Rabbit IgG H&L (HRP) (<a href='/en-us/products/secondary-antibodies/goat-rabbit-igg-h-l-hrp-ab97051'>ab97051</a>) at 1/100000 dilution

Predicted band size: 23 kDa

Observed band size: 28 kDa,32 kDa

false

Exposure time: 92s

• WB

Supplier Data

Western blot - Anti-CD69 antibody [EPR21814] (AB233396)

Blocking/Dilution buffer : 5% NFDM/TBST.

Exposure times : Lanes 1-5 and 7-8 : 3 minutes; Lane 6 : 26 seconds.

We observe an extra band at approximately 15 kDa that we do not yet have an explanation for; it could be non-specific.

This blot was developed using a higher sensitive ECL substrate.

All lanes:

Western blot - Anti-CD69 antibody [EPR21814] (ab233396) at 1/1000 dilution

Lane 1:

HL-60 (human Acute Promyelocytic Leukemia promyeloblast) whole cell lysate at 20 µg

Lane 2:

K562 (human chronic myelogenous leukemia lymphoblast) whole cell lysate at 20 µg

Lane 3:

U937 (human histiocytic lymphoma monocyte) whole cell lysate at 20 µg

Lane 4:

Human tonsil at 20 µg

Lane 5:

Human lymphoma at 20 µg

Lane 6:

H9 (human lymphoma T lymphocyte) whole cell lysate at 20 µg

Lane 7:

Human thymus at 20 µg

Lane 8:

Human colon at 20 µg

Secondary

All lanes:

Western blot - Goat Anti-Rabbit IgG H&L (HRP) (<a href='/en-us/products/secondary-antibodies/goat-rabbit-igg-h-l-hrp-ab97051'>ab97051</a>) at 1/100000 dilution

Predicted band size: 23 kDa

Observed band size: 24 kDa,32 kDa

false