To assess ASC potency, each ASC line was tested for its ability to inhibit lymphocyte proliferation using a mixed leukocyte reaction (MLR). Peripheral blood mononuclear cell (PBMC) isolation and MLRs were carried out exactly as described 1 . In brief, PBMCs were activated with 5 microgram concanavalin A (Con‐A; Sigma‐Aldrich), stained with a viability dye (Fixable Viability Dye eFlour780; eBioscience, San Diego, CA), proliferation was measured via BrdU incorporation (BrdU Flow Kit; BD Biosciences; San Jose, CA), and analyzed on a flow cytometer (Cytomics FC500). Flow cytometry data were analyzed using FlowJo flow cytometry software (Tree Star, Inc.).
Cytomics fc500
Manufactured by BD
Sourced in United States
The Cytomics FC500 is a flow cytometry system designed for cell analysis and sorting. It is capable of quantifying multiple parameters of individual cells within a heterogeneous population. The system provides data on characteristics such as size, granularity, and fluorescence intensity.
Automatically generated - may contain errors
5 protocols using cytomics fc500
1
Evaluating ASC Immunomodulatory Potential
2
CD4 Count and HCV RNA Quantification
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EDTA anticoagulated blood samples for CD4 count was by flow cytometry (BD Company, USA, CYTOMICS-FC500) at SHAPHC. In this study, we defined the baseline CD4 count as the last test performed within one month before DAA was initiated.
HCV RNA quantitative detection were performed at baseline, weeks 2, 8, and 12 during treatment, and at weeks 12 and 24 after the treatment. HCV RNA was measured with the COBAS TaqMan HCV test with a lower limit of detection of 40 IU/ml. Samples shown to be positive for the presence of HCV by reverse transcriptase- polymerase chain reaction (RT-PCR) testing were analyzed further to identify the HCV genotype (GT) using an immune line probe assay (INNO-LiPA, Bayer Diagnostics, US).
HCV RNA quantitative detection were performed at baseline, weeks 2, 8, and 12 during treatment, and at weeks 12 and 24 after the treatment. HCV RNA was measured with the COBAS TaqMan HCV test with a lower limit of detection of 40 IU/ml. Samples shown to be positive for the presence of HCV by reverse transcriptase- polymerase chain reaction (RT-PCR) testing were analyzed further to identify the HCV genotype (GT) using an immune line probe assay (INNO-LiPA, Bayer Diagnostics, US).
3
Apoptosis Analysis by Flow Cytometry
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After treatment with BTCI, cells were digested with trypsin, harvested, washed twice with ice-cold PBS, fixed with 75% ethanol at −20 °C overnight, washed again and then incubated with RNase A (25 μg/ml, Bio Basic Inc., Markham, Ontario, Canada) at 37 °C for 30 min. Cells were washed once with PBS and incubated with PI (50 μg/ml) for 30 min at room temperature in the dark. The cells were re-suspended in 500 μl PBS and subjected to flow cytometry on a Cytomics FC500 flow cytometer (FACScan; BD Biosciences). A sub G0/G1 population seen to the left of the G0/G1 peak represents DNA fragmentation caused by apoptosis.
4
Isolation and Characterization of Human MSCs
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Human MSCs were derived from umbilical cord according to the protocols reported before, 16, 23 MSCs were cultured in low-sugar Dulbecco's Modified Eagle medium containing 10% Fetal bovine serum. After 24 h, nonadherent cells were removed. MSCs were identified by a Cytomics FC 500 flow cytometer (BD Biosciences, San Jose, CA) of CD73, CD90, and CD105 (FITC-conjugated antibodies, Cambridge, Abcam, U.K.). The third passage MSCs were adsorb into the CHSSs in vitro.
5
CD4 Count Determination in ART Patients
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EDTA-anticoagulated blood samples were subjected to flow cytometry (BD Company, USA, CYTOMICS-FC500) at the Shanghai Public Health Clinical Center (SHAPHC) to determine the CD4 count. In this study, the baseline CD4 count was defined as the last count measured up to one month before raltegravir-based ART began.
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