Muscle sections (4 μm in thickness) taken from the left thigh were frozen in cold 2-methylbutane and were stained with anti-CD8a (53-6.7) and anti-CD4 (RM4-5) (BD Biosciences). Muscle sections (4 μm in thickness) taken from the right thigh were fixed in formalin, dehydrated, embedded in paraffin, and were then incubated with rat monoclonal antibodies specific for CD3 (CD3-12; Serotec, Oxford, UK), F4/80 (A3-1: Abcam, Cambridge, UK), and myeloperoxidase (MPO; NeoMarkers, Fremont, CA). Six inflammatory mononuclear cell foci in the serial sections were studied. Stained cells were counted in every focus under high magnification (400×) using a light microscope. The mean score was used for analysis. The stained sections were evaluated by two independent observers who reported comparable results.
Cd3 12
Manufactured by Bio-Rad
Sourced in United Kingdom, Germany
The CD3-12 is a laboratory equipment product manufactured by Bio-Rad. It is a device used for the detection and quantification of the CD3 protein, which is a key component of the T-cell receptor complex in the immune system. The CD3-12 utilizes specific antibodies and detection methods to identify and analyze the presence of CD3 in biological samples.
Automatically generated - may contain errors
Lab products found in correlation
3 3 diaminobenzidine substrate, by Agilent Technologies (2 mentions)
Histofine simple stain maxpo rat, by Nichirei Biosciences (2 mentions)
Anti cd8a 53 6.7, by BD (1 mentions)
Anti cd4 rm4 5, by BD (1 mentions)
Axio imager z1 microscope, by Zeiss (1 mentions)
Anti human cd20, by Thermo Fisher Scientific (1 mentions)
Fjk 16, by Thermo Fisher Scientific (1 mentions)
Luxol fast blue (lfb), by Merck Group (1 mentions)
M3 84, by BD (1 mentions)
Mib 1, by Agilent Technologies (1 mentions)
Streptavidin biotin detection system, by Biocare Medical (1 mentions)
7 protocols using cd3 12
1
Immune Cell Profiling in Muscle
2
Visualizing Germinal Center Markers
3
Histopathological Analysis of Neuroinflammation
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The cerebellum and spinal cord were collected and fixed in 4% paraformaldehyde and embedded in paraffin. Inflammation was graded on 4 µm thick H&E‐stained sections. Per mouse, between four and six sections of the spinal cord were analyzed. Immunohistochemistry was performed for CD45 (30‐F11, 1:5000; BD, Breda, the Netherlands), mac‐3 (M3/84, 1:30; BD), CD3 (CD3‐12, 1:100; AbD Serotec, Puchheim, Germany) and FoxP3 (1:50, FJK‐16s; eBioscience, San Diego, CA, USA). The spinal cord and cerebellum were stained with luxol fast blue (0.1% LFB; Sigma Aldrich, Saint Louis, MO, USA) for observation of myelin loss. All stained sections were analyzed by an observer who was blinded to the experimental conditions.
4
Immunohistochemical CD3+ T-cell Staining
5
Immunohistochemical CD3+ T-cell Staining
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Paraffin-embedded sections were prepared from organs of treated mice. After antigen retrieval with citrate buffer (20 mM) in a pressure cooker (115 °), sections were incubated with a rat CD3 antibody (CD3-12, AbD Serotec) for 2 hr. After repeated washing, sections were incubated with Histofine simple stain MaxPo (Rat; Nichirei Biosciences) for 30 min. Staining was achieved by incubating with 3,3′-diaminobenzidine substrate (DAKO) and counterstaining with hematoxylin.
6
Immunophenotyping of Lymph Node Leukocytes
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Immunohistochemistry (IHC) was utilized to phenotype PLN leukocytes using antibodies for feline CD3 (Peter Moore, UC Davis, clone CD3-12), CD79 (AbD Serotec, clone HM57), CD204 (Trans Genic, clone SRA-E5), and Ki67 (Dako, clone MIB-1) on 4 micron serial sections with a streptavidin biotin detection system (Biocare Medical) and following the protocol described above, but using an anti-mouse secondary antibody. Substituting a matched mouse IgG for the primary antibody served as the negative control. The proportion of CD3+ T cells within each lymphoid follicle and Ki67+ cells in the paracortical zones were quantified utilizing ImageJ software (National Institutes of Health). Briefly, photomicrographs of lymphoid follicles and paracortical zones (five different follicles/cat and three random paracortical regions/cat) were obtained. Using ImageJ, the image threshold was adjusted to black (IHC positive cells) and white to minimize non-specific background. IHC positive cells were quantified by specifying the lower limit of cell size (100 pixels) and accounting for overlapping cells. The image was then assessed for total number of nuclei using the image-based tool for counting nuclei (ITCN). The frequency of IHC positive cells was expressed as a division of the number of positive cells divided by total number of nuclei.
7
CSF Analysis for Lymphoma Diagnosis
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The CSF was obtained from the cisterna magna for routine analysis. Erythrocyte and total nucleated cells counts were determined with a hemocytometer. Cytology preparations were prepared by centrifugation (Cytospin3; ThermoShandon, Pittsburgh, PA) of 500 mL of CSF for 5 minutes at 1000 rpm and routinely stained with Wright-Giemsa. A differential cell count was performed on 100 cells. Lymphoma was diagnosed if the cells consisted of a relatively monomorphic population of large (nuclei >2 times the diameter of a red blood cell) lymphocytes of immature appearance. In some instances, lymphoma was diagnosed on CSF examination, additional cytospin slides were used for immunophenotyping using the B-cell markers CD21 (CA2.1D6; P. F. Moore, UC Davis, CA) and CD79a (HM57; Dako Corp, Carpinteria, CA) and the T-cell markers CD3 (CD3-12; Serotec, Oxford, UK), CD4 (CA13.1E4; P. F. Moore), and CD8a (CA9.JD3; P. F. Moore).
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