Anti-PD1 抗体 [NAT105]

• IHC-Fr

Supplier Data

Immunohistochemistry (Frozen sections) - Anti-PD1 antibody [NAT105] (AB52587)

Immunohistochemical analysis of permeabilized frozen Human tonsil tissue labeling PD1 with ab52587 at 1/10000 dilution, followed by LeicaDS9800 (Bond™ PolymerRefine Detection). Counterstained with hematoxylin. The immunostaining was performed on a Leica Biosystems BOND® RX instrument.

• IHC-Fr

Supplier Data

Immunohistochemistry (Frozen sections) - Anti-PD1 antibody [NAT105] (AB52587)

Immunohistochemical analysis of permeabilized frozen Human cardiac muslce tissue labeling PD1 with ab52587 at 1/10000 dilution, followed by LeicaDS9800 (Bond™ PolymerRefine Detection). No staining on human cardiac muscle. Counterstained with hematoxylin. The immunostaining was performed on a Leica Biosystems BOND® RX instrument.

• mIHC

Collaborator

Multiplex immunohistochemistry - Anti-PD1 antibody [NAT105] (AB52587)

10-color fluorescence multiplex immunohistochemical analysis of human lung cancer tissue (formalin-fixed paraffin-embedded section).

Merged staining of anti-FOXP3 (ab215206; Cyan; TG540N), anti-PD1 (ab52587; Red; TG700N), anti-CD163 (ab182422; Brown; TG650N), anti-HLA-DR (ab92511; Yellow; TG570N), anti-CD4 (ab133616; Violet; TG620N), anti-CD8 alpha (ab101500; Purple; TG540S), anti-CD20 (ab9475; Grey; TG660S), anti-CD68 (ab192847; Green; TG520N), anti-Cytokeratin 19 (ab52625; Light blue; TG440N). TG470SN (dark blue) was used as a nuclear counter stain. The inset image shows the separate CD68 signal.

The section was incubated in nine rounds of staining; in the order of ab215206 (1/100 dilution), ab52587 (1/200 dilution), ab182422 (1/300 dilution), ab92511 (1/200 dilution), ab133616 (1/600 dilution), ab101500 (1/300 dilution), ab9475 (1/100 dilution), ab192847 (1/300 dilution), ab52625 (1/400 dilution); each using a separate fluorescent tyramide signal amplification system.

Sodium citrate antigen retrieval (pH6.0) was used in between rounds of tyramide signal amplification to remove the antibody from the previous round, to avoid any cross-reactivity.

Image acquisition was performed with TissueFAXS Spectra (TissueGnostics).

This image is courtesy of TissueGnostics Asia Pacific Limited

• Flow Cyt

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Flow Cytometry - Anti-PD1 antibody [NAT105] (AB52587)

Cells were fixed with 100µl 4% PFA for 10 min at RT, then permeabilized with 100µl 90% methanol for 30 min (-20° C) followed by blocking with 10% goat serum for 1h at room temperature. Incubate ab52587 for 30 min at room temperature (RT) and then incubate Goat anti-Mouse IgG (Alexa Fluor^® 488, ab150113) for 30 min at room temperature. Positive staining on MOLT-4 treated with 10ng/ml PMA and 500ng/ml Ionomycin for 24h, but weak staining on untreated MOLT-4.

• IHC-P

Supplier Data

Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-PD1 antibody [NAT105] (AB52587)

Immunohistochemical analysis of paraffin-embedded Human cardiac muscle tissue labeling PD1 with ab52587 at 1/100 dilution, followed by a ready to use LeicaDS9800 (Bond™ Polymer Refine Detection). No staining on human cardiac muscle. The immunostaining was performed on a Leica Biosystems BOND® RX instrument. Counterstained with Hematoxylin. Heat mediated antigen retrieval was performed with Citrate buffer (pH 6.0, Epitope Retrieval Solution 1) for 20 mins.

• ICC/IF

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Immunocytochemistry/ Immunofluorescence - Anti-PD1 antibody [NAT105] (AB52587)

Immunofluorescent analysis of 4% Paraformaldehyde-fixed, 0.1% TritonX-100 permeabilized MOLT-4 cells labelling PD1 with ab52587 at 1/100 dilution, followed by Goat Anti-Mouse IgG H&L (Alexa Fluor^® 488) preadsorbed (ab150117) at 1/1000 dilution at RT for 45 min. Recombinant Alexa Fluor^® 594 Anti-beta Tubulin antibody [EPR16774] (ab206369) was used as a counterstain at 1/50 dilution and was co-incubated with ab52587 overnight at 4° C. Nucleus were visualized using DAPI. Confocal image showing membranous staining in MOLT-4 cells treated with 500 ng/ml Ionomycin and 10 ng/ml Phorbol-12-myristate-13-acetate (PMA) for 24 hours, and no staining in MOLT-4 cells.

• IHC-P

Supplier Data

Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-PD1 antibody [NAT105] (AB52587)

Immunohistochemical analysis of paraffin-embedded Human tonsil tissue labeling PD1 with ab52587 at 1/100 dilution, followed by a ready to use LeicaDS9800 (Bond™ Polymer Refine Detection). The immunostaining was performed on a Leica Biosystems BOND® RX instrument. Counterstained with Hematoxylin. Heat mediated antigen retrieval was performed with Citrate buffer (pH 6.0, Epitope Retrieval Solution 1) for 20 mins.

• WB

Supplier Data

Western blot - Anti-PD1 antibody [NAT105] (AB52587)

Blocking and diluting buffer and concentration : 5% NFDM/TBST.

All lanes:

Western blot - Anti-PD1 antibody [NAT105] (ab52587) at 1/1000 dilution

Lane 1:

Untreated MOLT-4 (human lymphoblastic leukemia T lymphoblast) whole cell lysate at 20 µg

Lane 2:

MOLT-4 treated with 500ng/ml Ionomycin and 10ng/ml Phorbol-12-myristate-13-acetate (PMA) for 24 hours, whole cell lysate at 20 µg

false

• IHC-P

CiteAb

Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-PD1 antibody [NAT105] (AB52587)

PD1 Immunohistochemistry using Anti-PD1 antibody [NAT105] ab52587. Publication image and figure legend from Kaewkangsadan, V., Verma, C., et al. 2016, J Immunol Res, PubMed 27777963.

PD-1+ T cells in the sections of LLABCs, using IHC staining, at 400x magnification. Briefly, heat-mediated antigen retrieval was performed using citrate buffer, pH 6 (20 mins). The sections were then incubated with MAbs to PD-1 (Abcam, ab52587) at a 1 : 100 dilution for 30 mins at RT. Polymeric HRP-linker antibody conjugate was used as secondary antibody. DAB chromogen was used to visualize the staining. The sections were counterstained with haematoxylin. (a) Low level of PD-1+ T cell infiltration; (b) high level of PD-1+ T cell infiltration. The average number of brown membrane-stained cells, regardless of intensity, in contact with tumour cells or within tumour cell nests (Itu : intratumoural) and in the interstitial stroma (Str : stromal) per HPF was counted.

• WB

CiteAb

Western blot - Anti-PD1 antibody [NAT105] (AB52587)

PD1 western blot using Anti-PD1 antibody [NAT105] ab52587. Publication image and figure legend from Zhao, Z., Shi, L., et al., 2018, Oncotarget, PubMed 29435173.

CTLs isolation and PD-1 knockout(A) The percentage of CD3+/CD8+ cells (CTLs) reached 95% after negative selection. (B) The CTLs were transduced with lentivirus co-expressing Cas9 and each of the indicated guide RNA. The expression of PD-1 guide RNA reduced the PD-1 expression in the transduced cells.

false