Five milliliters of CSF were obtained by lumbar spinal tap and centrifuged at 550g at 4°C for 5 minutes. The supernatant was removed, and the pellet was resuspended and incubated at 4°C for 20 minutes with fluorochrome-labeled antibodies against CD4, CD8, CD20, CD25, C-C chemokine receptor type 6 (CCR6), HLA-DR, CD14, and CD45 (BD Pharmingen Mouse Anti-Human Antibodies: CD4 PerCP, CD8 FITC, CD 20 Alexa Fluor 700, CD25 PE-Cy7, CD196 (CCR6) PE, Anti-HLA-DR V450, and CD14 APC; eBioscience mouse Anti-Human Antibody: CD45 EF 605). Cells were washed and resuspended in 200 μL of fluorescence-activated cell sorting (FACS) buffer before FACS analysis on a FACSCanto-II. Analysis of cell subset counts was performed with FlowJo. All CSF and serum tests were performed with researchers blinded to the clinical diagnosis and MRI results. CSF samples were measured by flow cytometry in a time frame of 90 minutes after lumbar spinal tab. CSF supernatants were stored in polypropylene tubes at −80°C until batched analysis of NfL and CHI3L1 concentrations.
Mouse anti human antibodies
Manufactured by BD
Sourced in United States
Mouse anti-human antibodies are laboratory reagents used in various immunological applications. They are designed to recognize and bind to specific human proteins or antigens, facilitating the detection and analysis of these targets in biological samples. These antibodies are a commonly used tool in research, diagnostic, and therapeutic settings.
Automatically generated - may contain errors
Lab products found in correlation
Cd3e hilyte 750 allophycocyanin h7apc, by BD (2 mentions)
Cd4 brilliant blue, by BD (2 mentions)
Cd8a phycoerythrin, by BD (2 mentions)
Cd14 brilliant violet, by BD (2 mentions)
Cd11c cyanin5 pe, by BD (2 mentions)
Cd2 brilliant ultraviolet, by BD (2 mentions)
Cd8 fitc, by Thermo Fisher Scientific (1 mentions)
Cd25 pe cy7, by Thermo Fisher Scientific (1 mentions)
Cd14 apc, by Thermo Fisher Scientific (1 mentions)
Cd4 percp, by Thermo Fisher Scientific (1 mentions)
Facscalibur, by BD (1 mentions)
Hla dr pe, by BD (1 mentions)
Tlr4 pe, by BD (1 mentions)
Annexin 5 fitc apoptosis detection kit, by Keygen Biotech (1 mentions)
Phosflow perm buffer 3, by BD (1 mentions)
Accuri c6 flow cytometer, by BD (1 mentions)
7 protocols using mouse anti human antibodies
1
CSF Immune Cell Profiling by Flow Cytometry
2
Flow Cytometry Analysis of Hematopoietic Cells
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A total of 1 × 106 cells were stained with mouse anti‐human antibodies (BD) for 30 minutes at 4°C in the dark and analysed with flow cytometry (FACSCalibur, BD) to determine the cell subpopulation in culture. These antibodies included PE‐CD34, BV421‐CD34, FITC‐CD38, APC‐CD38, BV510‐CD45A, PerCP‐Cy5.5‐CD90 and PE‐CD49f. According to the proportions in total cells, the numbers of CD34+ cells and CD34+CD38− cells were calculated, and the fold expansion was calculated using the numbers divided by the numbers on day 0.
3
Comprehensive Immune Cell Phenotyping
4
Quantifying Immune Markers in Myoblasts
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To quantify ICAM-1, HLA-DR, and TLR4 expression, human myoblasts were cultured for 48 hours in GM containing or not 250 µM of Co 2+ , Cr 3+ , and Co 2+ plus Cr 3+ . This experiment was also performed with the addition of human monocytes in GM (initial cell ratio at seeding 1:1). Cells were trypsinized, washed two times in PBS and incubated for 30 minutes on ice with one of the following mouse anti-human antibodies (all from BD Biosciences): CD54-PE (ICAM-1), HLA-DR-PE, or TLR4-PE. Negative controls were labeled with equivalent amounts of PE-labeled isotype-matched antibodies.
5
Comprehensive Immune Cell Phenotyping
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Cells were stained with the following mouse anti-human antibodies, purchased from BD Biosciences (USA): CD3e Hilyte 750 Allophycocyanin (H7APC), CD4 Brilliant Blue (BB) 630, CD8a Phycoerythrin (PE), CD14 Brilliant Violet (BV) 750, CD19 BV570, CD11c Cyanin5 PE, CD2 Brilliant Ultraviolet (BUV) 805, and CD57 BB790. After washing cells in PBS and fixing in 0.5% paraformaldehyde, samples were acquired on a BD Symphony A5 flow cytometer and data was analyzed using FlowJo v9 (USA).
6
Multiparametric Phenotyping of Human PBMCs
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Human freshly-isolated PBMCs at 106/tube for each individual were stained with antibodies specific for T and B cells. Cells were then surface stained with a mixture of the following mouse anti-human antibodies (BD Biosciences): Hu CD3 PE HIT3a, Hu CD19 PE-Cy7 SJ25C1, Hu CD196 (CCR6) BV421 11A9, Hu CD183 APC 1C6/CXCR3, Hu CXCR5 (CD185) BV510 RF8B2 and Hu CD4 FITC RPA-T4. These samples were incubated at room temperature for 30 min, protected from the light. Cells were washed twice and fixed with 2% paraformaldehyde until acquisition on a BD LSRFortessa.
7
Analyzing Apoptosis and T Cell Markers in HBV-T Cell Co-culture
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After co-culture with NC-T cells, HBV-T cells or amygdalin treated HBV-T cells for 48 h, the HepG2.2.15 were harvested for apoptosis analysis. Quantification of apoptotic cells was performed with Annexin V-FITC apoptosis detection kit (Keygen, China). Briefly, the HepG2.2.15 cell samples were harvested with 0.25% trypsin without EDTA after 48 h of infection and then washed twice with ice-cold PBS and re-suspended in 500 μl binding buffer. Then cells were incubated with 5 μl Annexin V-FITC specific antibodies and 5 μl propidium iodide (PI) then incubated for 15–20 min in the dark and detected by BD Accuri C6 flow cytometer (BD, USA) with an excitation wavelength of Ex = 488 nm and an emission wavelength of Em = 530 nm.
For T cell surface markers detection, Fixed T cells were permeabilized by adding 1 ml of cold Phosflow Perm Buffer III (BD Biosciences) and incubating for 30 min on ice. Samples were then washed and incubated with the mouse anti-human antibodies (BD Biosciences) anti-CD3-PE-Cy5, −CD4-FITC, −CD8-FITC, and p-STAT3-PE at room temperature for 1 h. Then, the cells were washed and collected for flow cytometry analysis on a BD Accuri C6 flow cytometer (BD, USA). Each experiment was repeated three times in triplicate.
For T cell surface markers detection, Fixed T cells were permeabilized by adding 1 ml of cold Phosflow Perm Buffer III (BD Biosciences) and incubating for 30 min on ice. Samples were then washed and incubated with the mouse anti-human antibodies (BD Biosciences) anti-CD3-PE-Cy5, −CD4-FITC, −CD8-FITC, and p-STAT3-PE at room temperature for 1 h. Then, the cells were washed and collected for flow cytometry analysis on a BD Accuri C6 flow cytometer (BD, USA). Each experiment was repeated three times in triplicate.
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