重组Anti-HLA Class 1 ABC抗体[EPR22172]

• IHC-Fr

Lab

Immunohistochemistry (Frozen sections) - Anti-HLA Class 1 ABC antibody [EPR22172] (AB225636)

IHC image of HLA Class 1 ABC staining in a section of human normal frozen spleen* performed on a Leica Biosystems BOND® RX instrument using the standard protocol. The section was fixed in 10% paraformaldehyde (10 min) prior to staining. The section was incubated with ab225636, 0.05ugml, for 15 mins at room temperature and detected using an HRP conjugated compact polymer system. DAB was used as the chromogen. The section was then counterstained with haematoxylin and mounted with DPX. The inset secondary-only control image is taken from an identical assay without primary antibody. For other IHC staining systems (automated and non-automated) customers should optimize variable parameters such as antigen retrieval conditions, primary antibody concentration and antibody incubation times. *Tissue obtained from the Human Research Tissue Bank, supported by the NIHR Cambridge Biomedical Research Centre

• IHC-P

Supplier Data

Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-HLA Class 1 ABC antibody [EPR22172] (AB225636)

Immunohistochemical analysis of paraffin-embedded human colonic carcinoma tissue labeling HLAB with ab225636 at 1/4000 dilution, followed by a ready to use Goat Anti-Rabbit IgG H&L (HRP). Membranous and cytoplasmic staining on tumor cells of human colonic carcinoma (PMID : 26310568, PMID : 17014712) 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-HLA Class 1 ABC antibody [EPR22172] (AB225636)

Immunohistochemical analysis of paraffin-embedded human stomach tissue labeling HLAB with ab225636 at 1/4000 dilution, followed by a ready to use Goat Anti-Rabbit IgG H&L (HRP). Membranous and cytoplasmic staining on lymph nodule of human stomach (PMID : 25294906) 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-HLA Class 1 ABC antibody [EPR22172] (AB225636)

Immunohistochemical analysis of paraffin-embedded human liver tissue labeling HLAB with ab225636 at 1/4000 dilution, followed by a ready to use Goat Anti-Rabbit IgG H&L (HRP). Positive staining on hepatic sinus endothelium of human liver (PMID : 26963506) 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).

• ICC/IF

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Immunocytochemistry/ Immunofluorescence - Anti-HLA Class 1 ABC antibody [EPR22172] (AB225636)

Immunofluorescent analysis of 100% methanol-fixed THP-1 (human monocytic leukemia cell line) cells labeling HLAB with ab225636 at 1/250 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 THP-1 cells. The nuclear counterstain is DAPI (blue).
Tubulin is detected with ab195889 Anti-alpha Tubulin antibody [DM1A] - Microtubule Marker (Alexa Fluor^® 594) at 1/200 dilution (red).
The negative control is the secondary antibody only.

Methanol is recommended for fixation as weak signal was observed using 4% PFA.

• IHC-Fr

Lab

Immunohistochemistry (Frozen sections) - Anti-HLA Class 1 ABC antibody [EPR22172] (AB225636)

IHC image of HLA Class 1 ABC staining in a section of human normal frozen liver* performed on a Leica Biosystems BOND® RX instrument using the standard protocol. The section was fixed in 10% paraformaldehyde (10 min) prior to staining. The section was incubated with ab225636, 0.05ugml, for 15 mins at room temperature and detected using an HRP conjugated compact polymer system. DAB was used as the chromogen. The section was then counterstained with haematoxylin and mounted with DPX. The inset secondary-only control image is taken from an identical assay without primary antibody. For other IHC staining systems (automated and non-automated) customers should optimize variable parameters such as antigen retrieval conditions, primary antibody concentration and antibody incubation times. *Tissue obtained from the Human Research Tissue Bank, supported by the NIHR Cambridge Biomedical Research Centre

• Flow Cyt

Supplier Data

Flow Cytometry - Anti-HLA Class 1 ABC antibody [EPR22172] (AB225636)

Flow cytometric analysis of THP-1 (human monocytic leukemia cell line) cell line labeling HLAB with ab225636 at 1/600 (red) compared with a Rabbit monoclonal IgG (ab172730) (black) and an unlabeled control (cells without incubation with primary antibody and secondary antibody) (blue). Goat anti rabbit IgG (Alexa Fluor^® 488, ab150077), at 1/2000 dilution was used as the secondary antibody.

Gated on viable cells.

• IP

Supplier Data

Immunoprecipitation - Anti-HLA Class 1 ABC antibody [EPR22172] (AB225636)

HLAB was immunoprecipitated from 0.35 mg THP-1 (human monocytic leukemia cell line) whole cell lysate with ab225636 at 1/30 dilution. Western blot was performed from the immunoprecipitate using ab225636 at 1/500 dilution. VeriBlot for IP Detection Reagent (HRP) (ab131366), was used for detection at 1/1000 dilution.
Lane 1 : THP-1 whole cell lysate 10 μg (Input).
Lane 2 : ab225636 IP in THP-1 whole cell lysate.
Lane 3 : Rabbit monoclonal IgG (ab172730) instead of ab225636 in THP-1 whole cell lysate.
Blocking/Dilution buffer : 5% NFDM/TBST.
Exposure time : 3 seconds.

All lanes:

Immunoprecipitation - Anti-HLA Class 1 ABC antibody [EPR22172] (ab225636)

Predicted band size: 44 kDa

Observed band size: 40 kDa,43 kDa

false

• WB

Unknown

Western blot - Anti-HLA Class 1 ABC antibody [EPR22172] (AB225636)

This antibody has cross reaction with human HLA-A and HLA-C.# Recombinant human HLA-A protein was purchased from abnova company (H00003105-P01).

Recombinant human HLA-C protein was purchased from novusbio company (NBP2-23210).

All lanes:

Western blot - Anti-HLA Class 1 ABC antibody [EPR22172] (ab225636) at 1/1000 dilution

Lane 1:

Recombinant human HLA-A protein (aa24-aa365) 10ng

Lane 3:

Recombinant human HLA-C protein (aa25-aa308) 10ng

Secondary

All lanes:

Goat Anti-Rabbit IgG, (H+L), Peroxidase conjugated (<a href='/products/secondary-antibodies/goat-rabbit-igg-h-l-hrp-ab97051'>ab97051</a>) at 1/20000 dilution

false

• WB

Supplier Data

Western blot - Anti-HLA Class 1 ABC antibody [EPR22172] (AB225636)

Blocking/Dilution buffer : 5% NFDM/TBST.

Weak reactive bands of undetermined proteins are observed in addition to the target band.

All lanes:

Western blot - Anti-HLA Class 1 ABC antibody [EPR22172] (ab225636) at 1/1000 dilution

Lane 1:

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

Lane 2:

THP-1 (human monocytic leukemia cell line), whole cell lysate at 20 µg

Lane 3:

HL-60 (human promyelocytic leukemia cell line), whole cell lysate at 20 µg

Lane 4:

Jurkat (human T cell leukemia cell line from peripheral blood), whole cell lysate at 20 µg

Lane 5:

HepG2 (human liver hepatocellular carcinoma cell line), whole cell lysate at 20 µg

Lane 6:

TF-1 (human bone marrow erythroleukemia cell line), whole cell lysate at 20 µg

Lane 7:

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

Lane 8:

Human tonsil at 20 µg

Secondary

All lanes:

Western blot - Goat Anti-Rabbit IgG H&L (HRP) (<a href='/products/secondary-antibodies/goat-rabbit-igg-h-l-hrp-ab97051'>ab97051</a>) at 1/50000 dilution

Predicted band size: 40 kDa

Observed band size: 40 kDa

false

Exposure time: 26s

• ICC/IF

Supplier Data

Immunocytochemistry/ Immunofluorescence - Anti-HLA Class 1 ABC antibody [EPR22172] (AB225636)

Immunofluorescent analysis of 100% methanol-fixed Raji (human Burkitt's lymphoma cell line) cells labeling HLAB with ab225636 at 1/250 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 Raji cells. The nuclear counterstain is DAPI (blue).
Tubulin is detected with ab195889 Anti-alpha Tubulin antibody [DM1A] - Microtubule Marker (Alexa Fluor^® 594) at 1/200 dilution (red).
The negative control is the secondary antibody only.

Methanol is recommended for fixation as weak signal was observed using 4% PFA.

• Flow Cyt

Supplier Data

Flow Cytometry - Anti-HLA Class 1 ABC antibody [EPR22172] (AB225636)

Flow cytometric analysis of Raji (human Burkitt's lymphoma cell line) cells labeling HLAB with ab225636 at 1/600 (red) compared with a Rabbit monoclonal IgG (ab172730) (black) and an unlabeled control (cells without incubation with primary antibody and secondary antibody) (blue). Goat anti-rabbit IgG (Alexa Fluor^® 488, ab150077), at 1/2000 dilution was used as the secondary antibody.

Gated on viable cells.