752-9956-94

antibody from Invitrogen Antibodies

Targeting: CD74 DHLAG

Immunohistochemistry (Supportive data in Antibodypedia)

Antibody Data

Product number

752-9956-94

Provider

Invitrogen Antibodies

Product name

HLA-DR Monoclonal Antibody (LN3), Alexa Fluor™ Plus 488

Antibody type

Monoclonal

Antigen

Other

Description

Description: The LN3 antibody reacts with the human major histocompatibility complex (MHC) class II, HLA-DR. HLA-DR is expressed on the surface of human antigen presenting cells (APC) including B cells, monocytes, macrophages, DCs, and activated T cells. HLA-DR is a heterodimeric transmembrane protein composed of alpha and beta subunits and plays an important role in the presentation of peptides to CD4^+ T lymphocytes. Applications Reported: The LN3 antibody has been reported for use in flow cytometric analysis, and immunohistochemical staining. Immunohistochemistry can be performed using frozen and Bouin's, or formalin/paraffin, human tissues. Applications Tested: This LN3 antibody has been tested by immunohistochemistry of formalin-fixed paraffin embedded tissue using high pH antigen retrieval and can be used at 10 µg/mL. It is recommended that the antibody be carefully titrated for optimal performance in the assay of interest. Using conjugate solutions: Centrifuge the protein conjugate solution briefly in a microcentrifuge before use; add only the supernatant to the experiment. This step will help eliminate any protein aggregates that may have formed during storage, thereby reducing nonspecific background staining.

Reactivity

Human

Host

Mouse

Conjugate

Green dye

Isotype

IgG

Antibody clone number

LN3

Vial size

500 µL

Concentration

0.2 mg/mL

Storage

4°C, store in dark

Immunohistochemistry

Supportive validation

Submitted by

Invitrogen Antibodies (provider)

Main image

Experimental details

Immunohistochemical analysis of HLA-DR was performed on FFPE human tonsil tissue. To expose the target protein, HIER was performed on de-paraffinized sections using BOND Epitope Retrieval Solution 2 (pH 9) for 10 min, followed by a 5 min cool down, a 1 min wash with ddH2O, and then transferred to 1X PBS. Tissues were permeabilized with 0.1% Triton X-100 in 1X PBS for 30 min, washed three times in 1X PBS and blocked with 3% BSA/5% normal goat serum/1X PBS for 1 hr at RT. Tissues were then probed with HLA-DR Monoclonal Antibody (LN3), Alexa Fluor™ Plus 488 (Product # 752-9956-94) at 10 µg/mL (left) or with HLA-DR Monoclonal Antibody (LN3) (Product # MA5-11966) at 1 µg/mL (right), respectively, in blocking solution for 1 hr at RT in a humidified chamber. Tissues were washed twice for 5 min each in 1X PBST (0.05% Tween-20), then once for 5 min in 1X PBS. Detection of the unconjugated primary antibody was performed using Goat anti-Mouse IgG (H+L) Secondary Antibody, Alexa Fluor™ Plus 594 (Product # A32742) at a dilution of 1:2,000 in blocking solution for 1 hr at RT, followed by two 5 min washes in 1X PBST then one 5 min wash in 1X PBS, and then stained with DAPI (Product # 62247) at 1 µg/mL diluted in 1X PBS for 5 min at RT. The sections were washed twice in 1X PBS followed by a final rinse in ddH2O and were mounted with ProLong™ Glass Antifade Mountant (Product # P36982). Images were captured on EVOS™ M7000 Imaging System (Product # AMF7000) at 20X magnification.

Supportive validation

Submitted by

Invitrogen Antibodies (provider)

Main image

Experimental details

Immunohistochemical analysis of HLA-DR was performed on formalin-fixed paraffin-embedded human tonsil tissue. To expose the target protein, HIER was performed on de-paraffinized sections using BOND Epitope Retrieval Solution 2 (pH 9) for 10 min, followed by a 5 min cool down, a 1 min wash with ddH2O, and then transferred to 1X PBS. Tissues were permeabilized with 0.1% Triton X-100 in 1X PBS for 30 min, washed three times in 1X PBS and blocked with 3% BSA/5% normal goat serum/1X PBS for 1 hr at RT. Following the removal of the blocking solution, tissues were probed with (left) or without (right) HLA-DR Monoclonal Antibody (LN3), Alexa Fluor™ Plus 488 (Product # 752-9956-94) at 10 µg/mL in blocking solution for 1 hr at RT in a humidified chamber. Tissues were then washed twice for 5 min each, in 1X PBS + 0.05% Tween-20 (PBST), and once for 5 min in 1X PBS. Tissues were then stained with DAPI (Product # 62247) at 1 µg/mL diluted in 1X PBS for 5 min at RT. The sections were washed twice in 1X PBS followed by a final rinse in ddH2O and were mounted with ProLong™ Glass Antifade Mountant (Product # P36982). Images were captured on EVOS™ M7000 Imaging System (Product # AMF7000) at 20X magnification.