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Cd68 kp1

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CD68 (KP1) is a widely used marker for macrophages in immunohistochemistry. It is a glycoprotein expressed by human monocytes and macrophages.

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Lab products found in correlation

Cd8 c8 144b, by Agilent Technologies (4 mentions) Background sniper, by Biocare Medical (2 mentions) H550s microscope, by Nikon (2 mentions) Diva decloaker, by Biocare Medical (2 mentions) Mach3 mouse hrp polymer detection, by Biocare Medical (2 mentions) Cd45 lca 2b11 pd7 26, by Agilent Technologies (2 mentions) Cd3 sp7, by Thermo Fisher Scientific (2 mentions) Cd20 l26, by Biocare Medical (2 mentions) Cd20 l26, by Agilent Technologies (2 mentions) Cd4 4b12, by Agilent Technologies (2 mentions) Autostainer link 48, by Agilent Technologies (2 mentions) Bx51 microscope, by Olympus (1 mentions) D2 40, by Abcam (1 mentions) Prolong gold, by Thermo Fisher Scientific (1 mentions) Ab34164, by Abcam (1 mentions) Vectra 3.0 multispectral imaging system, by PerkinElmer (1 mentions) Cd20 l26, by Leica (1 mentions) Cd3 f7, by Agilent Technologies (1 mentions) Opal 7 color ihc kit, by PerkinElmer (1 mentions) Cd3 polyclonal, by Agilent Technologies (1 mentions)

Close spelling variants of this product

Anti-CD68 FITC (clone KP1, CD68 (clone KP1, Anti‐CD68 antibody (clone KP1, Anti-human CD68 (clone KP1, Anti-CD68 (KP1, Mouse anti-CD68 (clone KP1,

12 protocols using cd68 kp1

1

Quantifying Renal Inflammatory Cells

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Formalin-fixed renal biopsy tissue was embedded in paraffin, mounted onto slides, and stained for myeloperoxidase (MPO) (rabbit-polyclonal, Abcam), and CD68 (KP1, DAKO-Agilent). The HIER method was used for antigen retrieval (BioGenex, San Ramon, CA). Images from 12 random fields within each slide were acquired at 400× magnification using an Olympus BX51 microscope (Olympus, Tokyo, Japan) and processed using ImageJ software (NIH, Bethesda, MD). Cell counts for each image were quantified using color-separation and background-subtraction, automatic thresholding (e.g., Maximum-Entropy, Triangle, Huang), and particle-analysis algorithms.
Zitur L.J., Chlebeck P.J., Odorico S.K., Danobeitia J.S., Zens T.J., Van Kooten C., Eerhart M., Reyes J.A., Springer M.L., Coonen J.M., Brunner K.G., Capuano S.V., D’Alessandro A.M, & Fernandez L.A. (2019). Brain Death Enhances Activation of the Innate Immune System and Leads to Reduced Renal Metabolic Gene Expression. Transplantation, 103(9), 1821-1833.
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2

Multiplex Immunofluorescence Staining for Cell Markers

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Multiplex staining was performed following the Opal Tyramide amplification signal protocol for the markers D2–40 (D2/40; Abcam, Cambridge, MA, USA) labeled with fluorophore 520 (Perkin Elmer, Akron, OH, USA); CD68 (KP1; Agilent, Santa Clara, CA, USA) labeled with fluorophore 520; and CD206 (5C11; LSBio, Seattle, WA, USA) labeled with fluorophore 570 (Perkin Elmer). Nuclei were stained with 4′,6-diamidino-2-phenylindole (DAPI; Perkin Elmer) and cover slips were applied to the slides with ProLong Gold (Life Technologies, Grand Island, NY, USA). The markers were distributed between two different panels of three colors each. The first panel included D2–40 and CD206; the second, CD68 and CD206. Slides containing prostate sections were deparaffinized in xylene and rehydrated as described above. For antigen retrieval, slides were microwaved with citrate H25 pH 6.0 buffer at 100 W for 1 min and 20 W for 15 min. After cooling for 15 min, they were treated with dual horseradish peroxidase block for 10 min, and the primary and secondary antibodies and fluorophores were applied according to their respective dilutions and incubation periods in a humid chamber. For the second round of primary antibody application, slides were first subjected to the same microwave antigen retrieval steps described above.
Zarif J.C., Baena-Del Valle J.A., Hicks J.L., Heaphy C.M., Vidal I., Luo J., Lotan T.L., Hooper J.E., Isaacs W.B., Pienta K.J, & De Marzo A.M. (2018). Mannose Receptor–positive Macrophage Infiltration Correlates with Prostate Cancer Onset and Metastatic Castration-resistant Disease. European urology oncology, 2(4), 429-436.
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3

Immunohistochemical Analysis of Immune Cells

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We performed immunohistochemical analysis of target antigen in the serial section of biopsies using the following antibodies and dilutions: (1) CD68 for macrophages (Mφ) in intact tissues. CD68 (KP1), a mouse monoclonal antibody (1:50) was derived from Dako. CD68 antigen (clone KP1), which we used for our current study as a marker of matured and activated Mφ, is a glycosylated trans‐membrane glycoprotein that is mainly located in lysosomes. (2) Anti‐Syndecan‐1 antibody [1:200, mouse monoclonal (B‐A38) to Syndecan (CD138), ab34164, Abcam] was used to immunolocalize plasma cells. (3) Non‐immune immunoglobulin (Ig) G1 (1:50), a mouse monoclonal antibody from Dako, was used as a negative control.
Khan K.N., Fujishita A., Ogawa K., Koshiba A., Mori T., Itoh K., Nakashima M, & Kitawaki J. (2021). Occurrence of chronic endometritis in different types of human adenomyosis. Reproductive Medicine and Biology, 21(1), e12421.
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4

Immunohistochemical Analysis of Human Cells

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Tissues for IHC were harvested from MoM and fixed in 4% paraformaldehyde for 24 hour at 4°C, embedded in paraffin, cut into 5-μm sections and mounted onto poly-L-lysine coated glass slides. Following paraffin removal, antigen retrieval (DIVA Decloaker, Biocare Medical, Cat. DV2004.) and blocking of non-specific Ig-binding sites (Background Sniper, Biocare Medical), tissue sections were stained with primary antibodies overnight at 4°C and developed with a biotin-free horseradish peroxidase (HRP)-polymer system (MACH3 Mouse HRP-Polymer Detection, Biocare Medical). All tissue sections were then counterstained with hematoxylin. Primary antibodies directed against CD45 LCA (2B11&PD7/26, Dako), CD3 (SP7, Thermo Scientific), CD20 (L26, Biocare Medical) and CD68 (KP1, Dako) were used to identify human cells in the spleen, liver and lung. For comparison, tissue sections were stained with mouse IgG1k or IgG2a isotype controls. Light microscopy images were taken with a Nikon H550S microscope at 40×.
Honeycutt J.B., Thayer W.O., Baker C.E., Ribeiro R.M., Lada S.M., Cao Y., Cleary R.A., Hudgens M.G., Richman D.D, & Garcia J.V. (2017). HIV persistence in tissue macrophages of humanized myeloid only mice during antiretroviral therapy. Nature medicine, 23(5), 638-643.
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5

Immunohistochemical Analysis of Human Cells

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Tissues for IHC were harvested from MoM and fixed in 4% paraformaldehyde for 24 hour at 4°C, embedded in paraffin, cut into 5-μm sections and mounted onto poly-L-lysine coated glass slides. Following paraffin removal, antigen retrieval (DIVA Decloaker, Biocare Medical, Cat. DV2004.) and blocking of non-specific Ig-binding sites (Background Sniper, Biocare Medical), tissue sections were stained with primary antibodies overnight at 4°C and developed with a biotin-free horseradish peroxidase (HRP)-polymer system (MACH3 Mouse HRP-Polymer Detection, Biocare Medical). All tissue sections were then counterstained with hematoxylin. Primary antibodies directed against CD45 LCA (2B11&PD7/26, Dako), CD3 (SP7, Thermo Scientific), CD20 (L26, Biocare Medical) and CD68 (KP1, Dako) were used to identify human cells in the spleen, liver and lung. For comparison, tissue sections were stained with mouse IgG1k or IgG2a isotype controls. Light microscopy images were taken with a Nikon H550S microscope at 40×.
Honeycutt J.B., Thayer W.O., Baker C.E., Ribeiro R.M., Lada S.M., Cao Y., Cleary R.A., Hudgens M.G., Richman D.D, & Garcia J.V. (2017). HIV persistence in tissue macrophages of humanized myeloid only mice during antiretroviral therapy. Nature medicine, 23(5), 638-643.
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6

Multiplex Immunofluorescence Analysis of NSCLC and Brain Metastases

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As described in a previous report [25 ], multiplex immunofluorescence staining was performed using the Opal 7-Color IHC Kit (PerkinElmer, Waltham, MA, USA) in the FFPE tissue sections of a typically paired NSCLC and its brain metastases. The stained slides were scanned by a Vectra 3.0 multispectral imaging system (PerkinElmer, Waltham, MA, USA). The immunofluorescence markers consisted of B7-H4 (1:150 dilution, clone D1M8I, Cell Signaling Technology), CD3 (F7.2.38, prediluted, Dako), CD8 (C8/144B, prediluted, Dako), CD20 (L26, ready-to-use, Leica), CD68 (KP1, prediluted, Dako), and CK (AE1/AE3, prediluted, Dako). DAPI was used for nuclei highlighting. After dewaxing and rehydration, antigen retrievals were performed using a Meidi microwave (Meidi, China). For each primary antibody, tyramide signal amplification linked to specific fluorochrome from the multiplex immunofluorescence staining Kit was used for incubation and visualization. We performed the complete multiplex immunofluorescence staining procedure following the manufacturer’s instructions. In addition, human tonsil FFPE tissues were analyzed with and without primary antibodies following the same multiplex immunofluorescence staining procedure to establish positive and negative (autofluorescence) controls.
Liu J.S., Cai Y.X., He Y.Z., Xu J., Tian S.F, & Li Z.Q. (2024). Spatial and temporal heterogeneity of tumor immune microenvironment between primary tumor and brain metastases in NSCLC. BMC Cancer, 24, 123.
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7

Immunohistochemical Analysis of Tissue Samples

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We reviewed hematoxylin-eosin stained slides prepared from formalin-fixed, paraffin-embedded tissue sections. In cases with bone marrow involvement, Wright-Giemsa–stained aspirate smears, touch imprints, and hematoxylin-eosin stained slides of aspirate clot and biopsy specimens were reviewed.
Immunohistochemical analysis was performed using 4-mm-thick formalin-fixed, paraffin-embedded tissue sections using heat-induced antigen retrieval, an avidin biotin-peroxidase complex method and an automated immunostainer (Ventana Biotech, Tucson, AZ, USA). A variable panel of antibodies was used as appropriate for establishing the diagnosis as described elsewhere.12 (link) The antibodies used were reactive with the following antigens: CD3 (1:100) (Lab Vision; Thermo Fisher Scientifics, Fremont, CA); CD20 (1:200), CD68 (KP-1; 1:900;), CD138 (M1-15; 1:600), IgG (1:40000), IgM (1:3000), IgA (1:5000), kappa (1:20,000) and lambda (1:20,000) (Dako North America, Inc., Carpenteria, CA). Detection was performed with biotinylated secondary antibody, horseradish peroxidase and 3,3’-diaminobenzidine as a chromogen. Slides were counterstained with haematoxylin. Appropriate negative and positive control slides were run in parallel.
Kanagal-Shamanna R., Xu-Monette Z.Y., Miranda R.N., Dogan A., Zou D., Luthra R., Weber D., O’Malley D.P., Jorgensen J.L., Khoury J.D., Bueso-Ramos C.E., Orlowski R.Z., Medeiros L.J, & Young K.H. (2015). Crystal-Storing Histiocytosis: A Clinicopathologic Study of 13 Cases. Histopathology, 68(4), 482-491.
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8

Immunohistochemical Analysis of Tumor Markers

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Immunohistochemistry was performed on 2‐μm FFPE sections in an automated immunostainer (Autostainer Link48; Dako, Glostrup, Denmark) according to the manufacturer's instructions. The following antibodies were used: β‐catenin 14 (1 : 1000; BD Transduction, South San Francisco, CA, USA), CD3 polyclonal (1 : 400; Dako), CD4 4B12 (1 : 300; Dako), CD8 C8/144B (1 : 20; Dako), CD20 L26 (1 : 400; Dako), CD21 1F8 (1 : 50; Dako), CD56 123C3 (1 : 400; Dako), CD68 KP1 (1 : 3000; Dako), CD163 10D6 (1 : 200; Novocastra, Newcastle, UK), FOXP3 259D/C7 (1 : 200; BD Pharmingen), PD‐L1 SP142 (1 : 30; SPRING, Pleasanton, CA, USA), PD1 NAT105 (1 : 100; Biocare Medical, Pikenine, Concord, NC, USA), and EZH2 5246S (1 : 50; Cell Signaling, Leiden, Netherlands).
Colombo C., Belfiore A., Paielli N., De Cecco L., Canevari S., Laurini E., Fermeglia M., Pricl S., Verderio P., Bottelli S., Fiore M., Stacchiotti S., Palassini E., Gronchi A., Pilotti S, & Perrone F. (2017). β‐Catenin in desmoid‐type fibromatosis: deep insights into the role of T41A and S45F mutations on protein structure and gene expression. Molecular Oncology, 11(11), 1495-1507.
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9

Histopathological Analysis of Neurological Samples

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Formalin-fixed paraffin-embedded 5-µm thick sections were stained with haematoxylin and eosin (H&E), Luxol fast blue and periodic acid Schiff (LFB/PAS) for myelin, and modified Bielschowsky silver for axons. Immunohistochemistry was performed with the avidin–biotin-complex method,13 (link) using primary antibodies specific for GFAP (1:100, DAKO), neurofilament protein (1:800, steam antigen retrieval with citric acid buffer pH 6.0, DAKO), AQP4 (1:250, Sigma-Aldrich), myelin proteolipid protein (PLP) (1:500, Serotec), myelin-associated glycoprotein (MAG) (1:1000, Abcam), myelin-oligodendrocyte glycoprotein (MOG) (1:1000, Abcam), CD68 KP1 (1:100, DAKO), C9neo (monoclonal B7 and polyclonal, 1:200, from Professor Paul Morgan, Cardiff, UK) and Alpha-B-crystallin (1:1000, Millipore).
Guo Y., Lennon V.A., Parisi J.E., Popescu B., Vasquez C., Pittock S.J., Howe C.L, & Lucchinetti C.F. (2021). Spectrum of sublytic astrocytopathy in neuromyelitis optica. Brain, 145(4), 1379-1390.
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10

Multiplex Immunostaining of Immune Cells

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We carried out double staining on formalin‐fixed paraffin‐embedded sections. First, the 4‐μm‐thick sections were immunostained using anti‐BTLA antibody or anti‐Cbl‐b antibody as the primary antibody, and visualized with 3,3′‐diaminobenzidine. After the tissue sections had been treated with glycine–HCl (pH 2.5), they were subjected to immunofluorescence staining using antibodies against each of the following antigens: CD1a (O10, Lab Vision, Fremont, CA, USA), CD3, CD4, CD8, CD14 (7, Leica Microsystems), CD20 (L26, DAKO), CD56 (1B6, Leica Microsystems), CD68 (KP1, DAKO), CD207 (12D6, Leica Microsystems), CD208 (104.G4, Immunotech, Fullerton, CA, USA), FOXP3, BTLA, and Cbl‐b. Immunostained tissue sections were analyzed with a confocal microscope (LSM5 Pascal; Carl Zeiss, Jena, Germany) equipped with a 15‐mW Kr/Ar laser.
Oguro S., Ino Y., Shimada K., Hatanaka Y., Matsuno Y., Esaki M., Nara S., Kishi Y., Kosuge T, & Hiraoka N. (2015). Clinical significance of tumor‐infiltrating immune cells focusing on BTLA and Cbl‐b in patients with gallbladder cancer. Cancer Science, 106(12), 1750-1760.
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