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Cd8 hit8a

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The CD8 HIT8a is a laboratory equipment product. It is designed to perform a specific function related to CD8 cells. Additional details about its core function and intended use are not available.

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Lab products found in correlation

Cd14 clone m5e2, by BD (2 mentions) Cd163 clone ghi 61, by BD (2 mentions) Cd86 clone it2, by BD (2 mentions) Cd25 clone 2a3, by BD (2 mentions) Cd127 biotin clone hil 7r m21, by BD (2 mentions) Cd56 clone b159, by BD (2 mentions) 7aad dead exclusion stain, by BD (2 mentions) Cd3 clone hit3a, by BD (2 mentions) Cd4 clone rpa t4, by BD (2 mentions) Hla dr clone g46 6, by BD (2 mentions) Facscalibur, by BD (2 mentions) Fixable viability dye, by Thermo Fisher Scientific (1 mentions) Cd4 sk3, by BD (1 mentions) Anti epha2, by R&D Systems (1 mentions) Sj25c1, by BD (1 mentions) Anti mouse igg f ab 2 fragment specific antibody, by Jackson ImmunoResearch (1 mentions) Facs canto 2 instrument, by BD (1 mentions) J3 119, by Beckman Coulter (1 mentions) Phosphate buffered saline (pbs), by Lonza (1 mentions) Cd80 clone l307, by BD (1 mentions)

3 protocols using cd8 hit8a

1

Phenotypic Characterization of PBMCs

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Freshly isolated PBMCs were washed once in FACS buffer (PBS supplemented with 3% fetal bovine serum and 0.1% sodium azide) and stained for flow cytometry. Antibodies used; CD14 clone M5E2, HLA-DR clone G46-6, CD86 clone IT2.2, CD163 clone GHI/61, CD3 clone HIT3a, CD4 clone RPA-T4, CD8 HIT8a, CD25 clone 2A3, CD127-biotin clone HIL-7R-M21, CD56 clone B159, all from BD Biosciences. All analyzes were performed on a FACS Calibur and gated on viable PBMCs using 7AAD dead exclusion stain (BD Biosciences, San Jose, CA, USA).
Larsson A.M., Roxå A., Leandersson K, & Bergenfelz C. (2019). Impact of systemic therapy on circulating leukocyte populations in patients with metastatic breast cancer. Scientific Reports, 9, 13451.
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2

Comprehensive Flow Cytometry Analysis of CAR T-cells

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The FACSCanto II instrument (BD Biosciences) was used for acquisition and FlowJo v10 (FlowJo) was used for analysis of flow cytometry data. Samples were washed with and stained in PBS (Lonza) with 1% FBS. For all experiments, known negatives (e.g. NT T-cells) served as gating controls. eBioscience Fixable viability dyes (Invitrogen) were used to exclude dead cells from analysis. T-cells were evaluated for CAR expression 5-10 days post transduction using anti-CD19 (J3-119, Beckman Coulter; SJ25C1, BD Biosciences) or anti-human IgG F(ab')2 fragment specific antibody (Jackson ImmunoResearch) for EphA2-CAR. Anti-mouse IgG F(ab')2 fragment specific antibody (Jackson ImmunoResearch) was used to detect HER2-CAR expression. GM18 expression was analyzed by staining with anti-CD116 (4H1, BioLegend). CAR T-cell phenotype was established using the following antibodies: CD4 (SK3, BD Biosciences), CD8 (HIT8a, BD Biosciences), CD45RA (HI100, BD Biosciences), and CCR7 (G043H7, BioLegend; 150503, BD Biosciences). The beta chain of GM18 was detected with anti-CD131 (3D7, BD Biosciences). Cell surface EphA2 expression was detected using anti-EphA2 (371805, R&D Systems).
Lange S., Sand L.G., Bell M., Patil S.L., Langfitt D, & Gottschalk S. (2021). A chimeric GM-CSF/IL18 receptor to sustain CAR T-cell function. Cancer discovery, 11(7), 1661-1671.
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3

Flow Cytometric Analysis of PBMCs and Monocytes

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PBMCs (10 000–50 000 cells) and isolated monocytes (5000–10 000 cells) were immediately stained for flow cytometry for a total of 20 min at 4°C. Due to inadequate sample amount, we were not able to perform all flow cytometric analyses on all patients. Antibodies used; CD14 clone M5E2 (1:10), HLA-DR clone G46-6 (1:50), CD80 clone L307.4 (1:15), CD86 clone IT2.2 (1:15), CD83 clone HB15e (1:15), CD33 clone WM53 (1:10), CD163 clone GHI/61 (1:15), CD16 clone 3G8 (1:20), CD3 clone HIT3a (1:25), CD4 clone RPA-T4 (1:25), CD8 HIT8a (1:25), CD25 clone 2A3 (1:10), CD127-biotin clone HIL-7R-M21 (1:10), CD56 clone B159 (1:10), all from BD Biosciences. Cells were analyzed using a FACSCalibur (BD Biosciences, San Jose, CA, USA). Analyzes were performed gated on PBMCs (≥2000 events per sample) and using 7AAD dead exclusion stain (BD Biosciences). Blood dendritic cell analyzes were performed using Blood DC enumeration kit according the manufacturer’s instructions (Miltenyi Biotec, Bergisch Gladbach, Germany). For co-receptor expression, relative mean fluorescence intensity (MFI) was chosen to avoid any variability in antibody batches.
Bergenfelz C., Larsson A.M., von Stedingk K., Gruvberger-Saal S., Aaltonen K., Jansson S., Jernström H., Janols H., Wullt M., Bredberg A., Rydén L, & Leandersson K. (2015). Systemic Monocytic-MDSCs Are Generated from Monocytes and Correlate with Disease Progression in Breast Cancer Patients. PLoS ONE, 10(5), e0127028.
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