PD-1 Antibody [Unconjugated]

Western blot shows lysates of EL-4 mouse lymphoblast cell line and CTLL-2 mouse cytotoxic T cell line (negative control). PVDF membrane was probed with 0.2 µg/mL of Goat Anti-Mouse PD-1 Antigen Affinity-purified Polyclonal Antibody (Catalog # AF1021) followed by HRP-conjugated Anti-Goat IgG Secondary Antibody (Catalog # HAF017). A specific band was detected for PD-1 at approximately 55 kDa (as indicated). This experiment was conducted under reducing conditions and using Immunoblot Buffer Group 1.

Western blot shows lysates of 293T human embryonic kidney cell line mock transfected or transfected with mouse PD-1. PVDF membrane was probed with 0.2 µg/mL of Goat Anti-Mouse PD-1 Antigen Affinity-purified Polyclonal Antibody (Catalog # AF1021) followed by HRP-conjugated Anti-Goat IgG Secondary Antibody (Catalog # HAF017). A specific band was detected for PD-1 at approximately 75 kDa (as indicated). This experiment was conducted under reducing conditions and using Immunoblot Buffer Group 1.

PD‑1 was detected in perfusion fixed frozen sections of mouse thymus using 15 µg/mL Mouse PD‑1 Antigen Affinity-purified Polyclonal Antibody (Catalog # AF1021) overnight at 4 °C. Tissue was stained (red) and counterstained (green). View our protocol for Fluorescent IHC Staining of Frozen Tissue Sections.

PD-1 was detected in perfusion fixed frozen sections of mouse spleen using Goat Anti-Mouse PD-1 Antigen Affinity-purified Polyclonal Antibody (Catalog # AF1021) at 5 µg/mL overnight at 4 °C. Tissue was stained using the Anti-Goat HRP-DAB Cell & Tissue Staining Kit (brown; Catalog # CTS008) and counterstained with hematoxylin (blue). Specific staining was localized to cytoplasm in splenocytes. View our protocol for Chromogenic IHC Staining of Frozen Tissue Sections.

PD‑1 was detected in perfusion fixed paraffin-embedded sections of mouse thymus using Goat Anti-Mouse PD‑1 Antigen Affinity-purified Polyclonal Antibody (Catalog # AF1021) at 1.7 µg/mL for 1 hour at room temperature followed by incubation with the Anti-Goat IgG VisUCyte™ HRP Polymer Antibody (Catalog # VC004). Before incubation with the primary antibody, tissue was subjected to heat-induced epitope retrieval using Antigen Retrieval Reagent-Basic (Catalog # CTS013). Tissue was stained using DAB (brown) and counterstained with hematoxylin (blue). Specific staining was localized to cell membranes. View our protocol for IHC Staining with VisUCyte HRP Polymer Detection Reagents.

Expression and glycosylation of Siglec-1 counter-receptors on resting and activated Tregs.(A) Eight Siglec-1 counter receptors were randomly selected for cytometry analysis. All experiments were performed three times and similar results were observed. (B) Normalized mRNA counts of Siglec-1 counter receptors were obtained from Treg RNA-Seq data. The result is visualized in a volcano plot. Only a small subset of counter-receptors showed strongly increased gene expression following Treg activation. The counter-receptors selected for flow cytometry analysis are highlighted in red. (C) Western blot showing that CD48 had a higher molecular weight in activated Tregs. (D) Western blot of PD-1 affinity-purified from resting and activated Tregs. PD 1 showed a dramatic decrease of molecular weight after PNGase F digestion (D), PD-1 from activated Tregs had higher molecular weight (E) and following sialidase treatment, PD-1 from resting and activated Tregs migrated at a slightly reduced molecular weight indicating the presence of sialic acids (F). Image collected and cropped by CiteAb from the following open publication (//pubmed.ncbi.nlm.nih.gov/35224210), licensed under a CC-BY license. Not internally tested by R&D Systems.

Expression and glycosylation of Siglec-1 counter-receptors on resting and activated Tregs.(A) Eight Siglec-1 counter receptors were randomly selected for cytometry analysis. All experiments were performed three times and similar results were observed. (B) Normalized mRNA counts of Siglec-1 counter receptors were obtained from Treg RNA-Seq data. The result is visualized in a volcano plot. Only a small subset of counter-receptors showed strongly increased gene expression following Treg activation. The counter-receptors selected for flow cytometry analysis are highlighted in red. (C) Western blot showing that CD48 had a higher molecular weight in activated Tregs. (D) Western blot of PD-1 affinity-purified from resting and activated Tregs. PD 1 showed a dramatic decrease of molecular weight after PNGase F digestion (D), PD-1 from activated Tregs had higher molecular weight (E) and following sialidase treatment, PD-1 from resting and activated Tregs migrated at a slightly reduced molecular weight indicating the presence of sialic acids (F). Image collected and cropped by CiteAb from the following open publication (//pubmed.ncbi.nlm.nih.gov/35224210), licensed under a CC-BY license. Not internally tested by R&D Systems.

Expression and glycosylation of Siglec-1 counter-receptors on resting and activated Tregs.(A) Eight Siglec-1 counter receptors were randomly selected for cytometry analysis. All experiments were performed three times and similar results were observed. (B) Normalized mRNA counts of Siglec-1 counter receptors were obtained from Treg RNA-Seq data. The result is visualized in a volcano plot. Only a small subset of counter-receptors showed strongly increased gene expression following Treg activation. The counter-receptors selected for flow cytometry analysis are highlighted in red. (C) Western blot showing that CD48 had a higher molecular weight in activated Tregs. (D) Western blot of PD-1 affinity-purified from resting and activated Tregs. PD 1 showed a dramatic decrease of molecular weight after PNGase F digestion (D), PD-1 from activated Tregs had higher molecular weight (E) and following sialidase treatment, PD-1 from resting and activated Tregs migrated at a slightly reduced molecular weight indicating the presence of sialic acids (F). Image collected and cropped by CiteAb from the following open publication (//pubmed.ncbi.nlm.nih.gov/35224210), licensed under a CC-BY license. Not internally tested by R&D Systems.

• Reactivity: Mu
• Applications: WB, Flow, IHC, CyTOF-ready
• Clonality: Polyclonal
• Host: Goat
• Conjugate: Unconjugated
• Concentration: LYOPH

PD-1 Antibody [Unconjugated] Summary

• Immunogen: S. frugiperda insect ovarian cell line Sf 21-derived recombinant mouse PD-1
  Leu25-Gln167
  Accession # Q02242
• Specificity: Detects mouse PD-1 in direct ELISAs and Western blots. In direct ELISAs, approximately 20% cross-reactivity with recombinant human PD‑1 is observed.
• Source: N/A
• Isotype: IgG
• Clonality: Polyclonal
• Host: Goat
• Gene: PDCD1
• Purity Statement: Antigen Affinity-purified

Applications/Dilutions

• Dilutions:
  • CyTOF-ready
  • Flow Cytometry 0.25 ug/10^6 cells
  • Immunohistochemistry 5-15 ug/mL
  • Western Blot 0.2 ug/mL

Packaging, Storage & Formulations

• Storage: Use a manual defrost freezer and avoid repeated freeze-thaw cycles.
  • 12 months from date of receipt, -20 to -70 °C as supplied.
  • 1 month, 2 to 8 °C under sterile conditions after reconstitution.
  • 6 months, -20 to -70 °C under sterile conditions after reconstitution.
• Buffer: Lyophilized from a 0.2 μm filtered solution in PBS with Trehalose. *Small pack size (SP) is supplied either lyophilized or as a 0.2 µm filtered solution in PBS.
• Preservative: No Preservative
• Concentration: LYOPH
• Reconstitution Instructions: Reconstitute at 0.2 mg/mL in sterile PBS.

Notes

This product is produced by and ships from R&D Systems, Inc., a Bio-Techne brand.

Alternate Names for PD-1 Antibody [Unconjugated]

• CD279 antigen
• CD279
• hPD-1
• PD1
• PD-1
• PD1hPD-l
• PDCD1
• programmed cell death 1
• programmed cell death protein 1
• Protein PD-1
• SLEB2

Background

Programmed Death-1 (PD-1) is type I transmembrane protein belonging to the CD28/CTLA-4 family of immunoreceptors that mediate signals for regulating immune responses (1). Other members of this family include CD28, CTLA-4, and ICOS (2-4). PD-1 is most closely related to CTLA-4 and shares approximately 24% amino acid (aa) sequence identity. The mouse PD-1 gene encodes a 288 aa protein with a putative 20 aa signal peptide, a 149 aa extracellular region with one immunoglobulin-like V-type domain, a 21 aa transmembrane domain, and a 98 aa cytoplasmic region. The cytoplasmic tail contains two tyrosine residues that form the immunoreceptor tyrosine-based inhibitory motif (ITIM) and immunoreceptor tyrosine-based switch motif (ITSM) that are important in mediating PD-1 signaling. Mouse and human PD-1 share approximately 69% aa sequence identity. Two B7 family proteins, PD-L1 (also called B7-H1) and PD-L2, have been identified as PD-1 ligands (5, 6). PD-1 is expressed on activated T cells, B cells, myeloid cells, and on a subset of thymocytes. PD-1 deficient mice have a defect in peripheral tolerance and spontaneously develop autoimmune diseases. Binding of PD-1 to PD-L1 or PD-L2 results in the inhibition of TCR-mediated proliferation and cytokine production as well as BCR-mediated signaling. PD-1 likely has an inhibitory role in regulating immune responses (1-4).

1. Ishida, Y. et al. (1992) EMBO J. 11:3887.
2. Sharpe, A.H. and G.J. Freeman (2002) Nat. Rev. Immunol. 2:116.
3. Coyle, A. and J. Gutierrez-Ramos (2001) Nat. Immunol. 2:203.
4. Nishimura, H. and T. Honjo (2001) Trends in Immunol. 22:265.
5. Latchman Y. et al. (2001) Nature Immun. 2:261.
6. Tamura, H. et al. (2001) Blood 97:1809.

Limitations

This product is for research use only and is not approved for use in humans or in clinical diagnosis. Primary Antibodies are guaranteed for 1 year from date of receipt.

Publications for PD-1 Antibody (AF1021)(25)

Publications using AF1021

• Soon-Oh Hong, Joonsung Kim, Sungmin Lee, Jaeil Shin, Hwanjun Choi, Eunjin Lee et al. Transgenic viral expression of PH-20, IL-12, and sPD1-Fc enhances immune cell infiltration and anti-tumor efficacy of an oncolytic virus Mol Ther Oncolytics 2023-09-21 [PMID: 37727704] (Western Blot, Human)
• Corria-Osorio J, Carmona SJ, Stefanidis E, Andreatta M, Ortiz-Miranda Y, Muller T, Rota IA, Crespo I, Seijo B, Castro W, Jimenez-Luna C, Scarpellino L, Ronet C, Spill A, Lanitis E, Romero P, Luther SA, Irving M, Coukos G Orthogonal cytokine engineering enables novel synthetic effector states escaping canonical exhaustion in tumor-rejecting CD8+ T cells Nat Immunol. 2023; 24(5): 869–883. 2023-04-20 [PMID: 37081150] (ELISA Capture, Mouse)
• T Kaya, N Mattugini, L Liu, H Ji, L Cantuti-Ca, J Wu, M Schifferer, J Groh, R Martini, S Besson-Gir, S Kaji, A Liesz, O Gokce, M Simons CD8+ T cells induce interferon-responsive oligodendrocytes and microglia in white matter aging Nature Neuroscience, 2022-10-24;25(11):1446-1457. 2022-10-24 [PMID: 36280798] (IHC, Mouse)
• L Codarri De, V Nicolini, M Hashimoto, M Karagianni, PC Schwalie, L Lauener, EM Varypataki, M Richard, E Bommer, J Sam, S Joller, M Perro, F Cremasco, L Kunz, E Yanguez, T Hüsser, R Schlenker, M Mariani, V Tosevski, S Herter, M Bacac, I Waldhauer, S Colombetti, X Gueripel, S Wullschleg, M Tichet, D Hanahan, HT Kissick, S Leclair, A Freimoser-, S Seeber, V Teichgräbe, R Ahmed, C Klein, P Umaña PD-1-cis IL-2R agonism yields better effectors from stem-like CD8+ T cells Nature, 2022-09-28;610(7930):161-172. 2022-09-28 [PMID: 36171284] (IHC, Xenograft)
• C Martins, M Silva, E Rasbach, P Singh, Y Itoh, JB Williams, E Statham, A Meurer, DV Martinez, A Brandenbur, MV Heppt, SR Barthel, T Schatton Distinct antibody clones detect PD-1 checkpoint expression and block PD-L1 interactions on live murine melanoma cells Scientific Reports, 2022-07-21;12(1):12491. 2022-07-21 [PMID: 35864188] (Western Blot, Mouse)
• Gang Wu, Gavuthami Murugesan, Manjula Nagala, Alex McCraw, Stuart M. Haslam, Anne Dell et al. Activation of regulatory T cells triggers specific changes in glycosylation associated with Siglec-1-dependent inflammatory responses Wellcome Open Research 2021-06-01 [PMID: 35224210] (Western Blot, Mouse)
• Gyu Jin Lee, Yukyung Jun, Yoon Kyung Jeon, Daekee Lee, Sanghyuk Lee, Jaesang Kim Mice transgenic for human CTLA4-CD28 fusion gene show proliferation and transformation of ATLL-like and AITL-like T cells OncoImmunology 2022-12-31 [PMID: 35003894]
• Irene Tirado-Gonzalez, Arnaud Descot, Devona Soetopo, Aleksandra Nevmerzhitskaya, Alexander Schäffer, Ivan-Maximilano Kur et al. AXL Inhibition in Macrophages Stimulates Host-versus-Leukemia Immunity and Eradicates Naïve and Treatment-Resistant Leukemia Cancer Discovery 2021-11-01 [PMID: 34103328]
• M Dudek, D Pfister, S Donakonda, P Filpe, A Schneider, M Laschinger, D Hartmann, N Hüser, P Meiser, F Bayerl, D Inverso, J Wigger, M Sebode, R Öllinger, R Rad, S Hegenbarth, M Anton, A Guillot, A Bowman, D Heide, F Müller, P Ramadori, V Leone, C Garcia-Cac, T Gruber, G Seifert, AM Kabat, JP Malm, S Reider, M Effenberge, S Roth, AT Billeter, B Müller-Sti, EJ Pearce, F Koch-Nolte, R Käser, H Tilg, R Thimme, T Böttler, F Tacke, JF Dufour, D Haller, PJ Murray, R Heeren, D Zehn, JP Böttcher, M Heikenwäld, PA Knolle Auto-aggressive CXCR6+ CD8 T cells cause liver immune pathology in NASH Nature, 2021-03-24;0(0):. 2021-03-24 [PMID: 33762736] (IHC, Mouse)
• Zhiwu Tan, Mei Sum Chiu, Chi Wing Yan, Kwan Man, Zhiwei Chen Eliminating mesothelioma by AAV-vectored, PD1-based vaccination in the tumor microenvironment Molecular Therapy - Oncolytics 2021-03-26 [PMID: 33614918]
• Bergren W. Crute, Rachel Sheraden, Vanessa L. Ott, Isaac T. W. Harley, Andrew Getahun, John C. Cambier Inhibitory Receptor Trap: A Platform for Discovery of Inhibitory Receptors That Utilize Inositol Lipid and Phosphotyrosine Phosphatase Effectors Frontiers in Immunology 2020-10-21 [PMID: 33193438]
• Ben-Long Liu, Qi-Lai Cao, Xin Zhao, Hui-Zhu Liu, Yu-Qiu Zhang Inhibition of TRPV1 by SHP-1 in nociceptive primary sensory neurons is critical in PD-L1 analgesia JCI Insight 2020-10-15 [PMID: 32960817]
• Gyu Jin Lee, Yukyung Jun, Hae Yong Yoo, Yoon Kyung Jeon, Daekee Lee, Sanghyuk Lee et al. Angioimmunoblastic T-cell lymphoma-like lymphadenopathy in mice transgenic for human RHOA with p.Gly17Val mutation OncoImmunology 2020-09-14 [PMID: 32923110]
• T Lerbs, L Cui, ME King, T Chai, C Muscat, L Chung, R Brown, K Rieger, T Shibata, G Wernig CD47 prevents the elimination of diseased fibroblasts in scleroderma JCI Insight, 2020-08-20;5(16):. 2020-08-20 [PMID: 32814713] (ICC, IHC, Mouse)
• R Hoefflin, S Harlander, S Schäfer, P Metzger, F Kuo, D Schönenber, M Adlesic, A Peighambar, P Seidel, CY Chen, M Consenza-C, A Jud, B Lahrmann, N Grabe, D Heide, FM Uhl, TA Chan, J Duyster, R Zeiser, C Schell, M Heikenwald, O Schilling, AA Hakimi, M Boerries, IJ Frew HIF-1alpha and HIF-2alpha differently regulate tumour development and inflammation of clear cell renal cell carcinoma in mice Nat Commun, 2020-08-17;11(1):4111. 2020-08-17 [PMID: 32807776] (IHC, Mouse)
• Ganesh Rao, Khatri Latha, Martina Ott, Aria Sabbagh, Anantha Marisetty, Xiaoyang Ling et al. Anti–PD-1 Induces M1 Polarization in the Glioma Microenvironment and Exerts Therapeutic Efficacy in the Absence of CD8 Cytotoxic T Cells Clinical Cancer Research 2020-09-01 [PMID: 32554515]
• Yan Liu, Ziqi Yang, Peilong Lai, Zijing Huang, Xiaowei Sun, Tian Zhou et al. Bcl-6-directed follicular helper T cells promote vascular inflammatory injury in diabetic retinopathy Theranostics 2020-03-04 [PMID: 32226551] (Immunohistochemistry, Mouse)
• F Ritzmann, C Jungnickel, G Vella, A Kamyschnik, C Herr, D Li, MM Menger, A Angenendt, M Hoth, A Lis, R Bals, C Beisswenge IL-17C-mediated innate inflammation decreases the response to PD-1 blockade in a model of Kras-driven lung cancer Sci Rep, 2019-07-17;9(1):10353. 2019-07-17 [PMID: 31316109] (IHC-P, Mouse)
• WJ Lee, YJ Lee, ME Choi, KA Yun, CH Won, MW Lee, JH Choi, SE Chang Expression of lymphocyte-activating gene 3 and T-cell immunoreceptor with immunoglobulin and ITIM domains in cutaneous melanoma and their correlation with programmed cell death 1 expression in tumor-infiltrating lymphocytes J. Am. Acad. Dermatol., 2019-03-14;81(1):219-227. 2019-03-14 [PMID: 30880064] (IHC, Human)
• Lin J. R, Izar B, et al. Highly multiplexed immunofluorescence imaging of human tissues and tumors using t-CyCIF and conventional optical microscopes. Elife 7/11/2018 [PMID: 29993362]
  
  Details:
  Citation using the Alexa Fluor 488 format of this antibody.
• P Lamichhane, L Karyampudi, B Shreeder, J Krempski, D Bahr, J Daum, KR Kalli, EL Goode, MS Block, MJ Cannon, KL Knutson IL-10 release upon PD-1 blockade sustains immunosuppression in ovarian cancer Cancer Res., 2017-10-09;0(0):. 10/9/2017 [PMID: 28993412]
• R Ruscher, RL Kummer, YJ Lee, SC Jameson, KA Hogquist CD8?? intraepithelial lymphocytes arise from two main thymic precursors Nat. Immunol., 2017-05-22;0(0):. 2017-05-22 [PMID: 28530714] (IHC, Mouse)
• Guoning Guo, Hong Li, Dayan Cao, Yongwen Chen The development of endometrial hyperplasia in aged PD-1-deficient female mice Diagnostic Pathology 2014-12-01 [PMID: 24886298]
• Kasagi S, Kawano S, Okazaki T, Honjo T, Morinobu A, Hatachi S, Shimatani K, Tanaka Y, Minato N, Kumagai S Anti-programmed cell death 1 antibody reduces CD4+PD-1+ T cells and relieves the lupus-like nephritis of NZB/W F1 mice. J. Immunol., 2010-02-05;184(5):2337-47. 2010-02-05 [PMID: 20139271] (IHC-Fr, Mouse)
• Bishop KD, Harris JE, Mordes JP, Greiner DL, Rossini AA, Czech MP, Phillips NE Depletion of the programmed death-1 receptor completely reverses established clonal anergy in CD4(+) T lymphocytes via an interleukin-2-dependent mechanism. Cell. Immunol., 2009-02-23;256(1):86-91. 2009-02-23 [PMID: 19230866] (Western Blot, Mouse)

Review for PD-1 Antibody (AF1021) (1)

Average Rating: 5

(Based on 1 review)

We have 1 review tested in 1 species: Human.

reviewed by: Sachin Puri Flow Human 06/15/2017

Summary

• Application: Flow Cytometry
• Sample Tested: Human T cell,Melanoma tissue
• Species: Human

Bioinformatics

• Gene Symbol: PDCD1
• Uniprot:
  • Q02242()