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Optilyse c solution

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Optilyse C Solution is a lysis reagent designed for the preparation of samples for flow cytometry. It is used to lyse red blood cells and fix leukocytes, allowing for the analysis of white blood cell populations.

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Lab products found in correlation

Cell fix solution, by BD (3 mentions) Facscanto 2 flow cytometer, by BD (2 mentions) Recombinant human c5a, by R&D Systems (2 mentions) Cytofix, by BD (2 mentions) Dxflex flow cytometer, by Beckman Coulter (1 mentions) Phosphate buffered saline (pbs), by Beckman Coulter (1 mentions) Lp vortex mixer, by Thermo Fisher Scientific (1 mentions) Anti hla dr ecd, by Beckman Coulter (1 mentions) Cd14 pc7, by Beckman Coulter (1 mentions) Optilyse c lysing solution, by Beckman Coulter (1 mentions) Bovine serum albumin (bsa), by Merck Group (1 mentions) Ethylene diamine tetraacetic acid (edta), by Thermo Fisher Scientific (1 mentions) Fc500 flow cytometer, by Beckman Coulter (1 mentions) Anti cd45 apc, by BD (1 mentions) Pe conjugated streptavidin, by Southern Biotech (1 mentions) Anti cd3 percp cy5, by BD (1 mentions)

Spelling variants of this product

Optilyse C (32 citations) OptiLyse C Lysing Solution (7 citations) OptiLyse C Lysis Solution (5 citations) OptiLyse Solution (2 citations)

9 protocols using optilyse c solution

1

Avdoralimab Inhibits C5a-induced Neutrophil Activation

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Various concentrations of avdoralimab were added to the blood samples in culture-treated 96-well U-bottom plates, and incubated for 20 min at 37°C under an atmosphere containing 5% CO2. We then added 18 nM human recombinant C5a (R&D Systems) to the samples. Plates were incubated for 20 minutes at 37°C under an atmosphere containing 5% CO2. Samples were then stained for flow cytometry analysis with anti-CD16 FITC and anti-CD11b PE-Cy5 antibodies. Erythrocytes were lysed with Optilyse C solution (Beckman Coulter, A11895), according to the manufacturer’s protocol, and resuspended in CytoFix (BD Bioscience 554655) for fixation. Cells were then analyzed on a FACS Canto II flow cytometer (BD Biosciences) with FACS Diva software.
Carvelli J., Demaria O., Vély F., Batista L., Benmansour N.C., Fares J., Carpentier S., Thibult M.L., Morel A., Remark R., André P., Represa A., Piperoglou C., Cordier P.Y., Le Dault E., Guervilly C., Simeone P., Gainnier M., Morel Y., Ebbo M., Schleinitz N, & Vivier E. (2020). Association of COVID-19 inflammation with activation of the C5a-C5aR1 axis. Nature, 588(7836), 146-150.
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2

Avdoralimab Inhibits C5a-induced Neutrophil Activation

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Various concentrations of avdoralimab were added to the blood samples in culture-treated 96-well U-bottom plates, and incubated for 20 min at 37°C under an atmosphere containing 5% CO2. We then added 18 nM human recombinant C5a (R&D Systems) to the samples. Plates were incubated for 20 minutes at 37°C under an atmosphere containing 5% CO2. Samples were then stained for flow cytometry analysis with anti-CD16 FITC and anti-CD11b PE-Cy5 antibodies. Erythrocytes were lysed with Optilyse C solution (Beckman Coulter, A11895), according to the manufacturer’s protocol, and resuspended in CytoFix (BD Bioscience 554655) for fixation. Cells were then analyzed on a FACS Canto II flow cytometer (BD Biosciences) with FACS Diva software.
Carvelli J., Demaria O., Vély F., Batista L., Benmansour N.C., Fares J., Carpentier S., Thibult M.L., Morel A., Remark R., André P., Represa A., Piperoglou C., Cordier P.Y., Le Dault E., Guervilly C., Simeone P., Gainnier M., Morel Y., Ebbo M., Schleinitz N, & Vivier E. (2020). Association of COVID-19 inflammation with activation of the C5a-C5aR1 axis. Nature, 588(7836), 146-150.
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3

MDSC Quantification in Hepatocellular Carcinoma

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We collected 4 mL of fasting peripheral venous blood from all HCC patients before GSM–TACE, 10 days after surgery, and 30 days after surgery using heparin anticoagulation tubes. We then added 100 μL whole blood to a dry blank tube, lysed it with 500 μL OptiLyse C Lysing Solution (Beckman Coulter Diagnostics, Brea, California, US), vortexed the solution, and incubated the lysed blood at room temperature for 15 min to assure lysis was complete. Then we added 2 mL phosphate-buffered saline (PBS), and we vortexed and centrifuged the mixture at 300 g for 5 min. After centrifugation, we aspirated the supernatant, resuspended the cell pellet in 500 μL PBS, added 5 μL each of CD14, CD11b, and HLA-DR antibodies, and mixed the solution low speed for 5 s. Finally, we analyzed the samples on a flow cytometer.
2.2.2 Antibodies and laboratory equipment: To determine MDSC frequency, we performed multicolor fluorescence-activated cell sorting analysis using Beckman Coulter CytExpert software version 1.1. We also used a DxFLEX Flow Cytometer (Beckman Coulter), a Labofuge 400R Centrifuge (Thermo Fisher Scientific, Waltham, Massachusetts, US), a LP Vortex Mixer(Thermo Fisher Waltham, Massachusetts, US), and the following anti-human monoclonal antibodies: CD11b-APC-Alexa Fluor 750, CD14-PC7, anti–HLA-DR-ECD, OptiLyse C solution, and PBS (all Beckman Coulter).
Yue Y., Ren Z., Liu Y, & Zhang Y. (2019). Changes in the frequency of myeloid-derived suppressor cells after transarterial chemoembolization with gelatin sponge microparticles for hepatocellular carcinoma. Journal of Interventional Medicine, 2(1), 21-26.
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4

Multiparameter Flow Cytometry Immunophenotyping

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Blood collected into EDTA tubes was washed in PBS before staining with LiveDead (Thermo Fisher) according to the manufacturer’s instructions. Cells were incubated with mouse serum to saturate the Fc receptors, and were then incubated with the appropriate antibody cocktail. Red blood cells were lysed in Optilyse C Solution (Beckman Coulter), according to the manufacturer’s instructions. Cells were fixed in Cell Fix solution (BD), according to the manufacturer’s instructions. Data were acquired in an LSRFortessaX20 flow cytometer. The FCS3.0 files obtained were exported from BD FACSDiva software and imported into FlowJo v.10.5.2 (BD Biosciences). Automated compensation was calculated with FACSDiva software and single-stained compensation beads. This compensation matrix was analyzed in detail in FlowJo, by investigating the N-by-N view feature and the pairwise expression of all proteins stained in this study. Fluorescence minus one (FMO) experiments were run before this study, to facilitate optimization of the compensation matrix. We then adjusted the compensation matrix where necessary due to over- or under-compensation by the automatic algorithm.
Carvelli J., Demaria O., Vély F., Batista L., Benmansour N.C., Fares J., Carpentier S., Thibult M.L., Morel A., Remark R., André P., Represa A., Piperoglou C., Cordier P.Y., Le Dault E., Guervilly C., Simeone P., Gainnier M., Morel Y., Ebbo M., Schleinitz N, & Vivier E. (2020). Association of COVID-19 inflammation with activation of the C5a-C5aR1 axis. Nature, 588(7836), 146-150.
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5

Optimized Flow Cytometry Staining and Compensation

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Cells were incubated with mouse serum to saturate the Fc receptors, and then with the appropriate antibody cocktail diluted in staining buffer: 1XPBS (Gibco), 0.2% BSA (Sigma), 0.02% sodium azide (Prolabo) and 2 mM EDTA (Invitrogen Life Technologies). Red blood cells were lysed in Optilyse C Solution (Beckman Coulter) according to the manufacturer’s instructions. Cells were fixed in Cell Fix solution (BD) according to the manufacturer’s instructions. Data were acquired in an LSRFortessaX20 flow cytometer. The FCS3.0 files obtained were exported from BD FACSDiva software and imported into FlowJo v.10.5.2 (BD Biosciences). Automatic compensation was calculated with FACSDiva software and single-stained compensation beads. This compensation matrix was analyzed in detail in FlowJo, with the N-by-N view feature and by investigating the pairwise expression of all proteins stained in this study. Fluorescence minus one (FMO) experiments were run before this study, to facilitate optimization of the compensation matrix. We then adjusted the compensation matrix where necessary due to over or undercompensation by the automatic algorithm.
Demaria O., Gauthier L., Vetizou M., Blanchard Alvarez A., Vagne C., Habif G., Batista L., Baron W., Belaïd N., Girard-Madoux M., Cesari C., Caratini M., Bosco F., Benac O., Lopez J., Fenis A., Galluso J., Trichard S., Carrette B., Carrette F., Maguer A., Jaubert S., Sansaloni A., Letay-Drouet R., Kosthowa C., Lovera N., Dujardin A., Chanuc F., Le Van M., Bokobza S., Jarmuzynski N., Fos C., Gourdin N., Remark R., Lechevallier E., Fakhry N., Salas S., Deville J.L., Le Grand R., Bonnafous C., Vollmy L., Represa A., Carpentier S., Rossi B., Morel A., Cornen S., Perrot I., Morel Y, & Vivier E. (2022). Antitumor immunity induced by antibody-based natural killer cell engager therapeutics armed with not-alpha IL-2 variant. Cell Reports Medicine, 3(10), 100783.
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6

Characterization of NK (CD3-CD56dim) Cells

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NK (CD3−CD56dim) cells were characterised using 5 μl fluorescein isothiocyanate (FITC) anti-human CD3 and 5 μl phycoerythrin (PE) antihuman CD56, (Beckman Coulter, UK). On adding the labelled monoclonal antibodies (MAbs) a gentle vortex was applied for 5 s and the FACS tubes were left in the dark for 15 min at room temperature (RT). 500 μl of optilyse C solution (Beckman Coulter, UK) was added to induce complete lysis of red blood cells, vortexed and left for another 15 min at RT in the dark. 500 μl of phosphate buffered saline (PBS) was added to the FACS tubes to stop the lysis reaction between the optilyse C and the whole blood. The whole blood mixture was vortexed at RT. 100 μl of flow count-fluorsphere beads (Beckman Coulter, UK) was added prior to analysis on the flow cytometer (Beckman Coulter, FC500). Total events acquired were 150,000.
Verma C., Kaewkangsadan V., Eremin J.M., Cowley G.P., Ilyas M., El-Sheemy M.A, & Eremin O. (2015). Natural killer (NK) cell profiles in blood and tumour in women with large and locally advanced breast cancer (LLABC) and their contribution to a pathological complete response (PCR) in the tumour following neoadjuvant chemotherapy (NAC): differential restoration of blood profiles by NAC and surgery. Journal of Translational Medicine, 13, 180.
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7

Immune Cell Profiling using Flow Cytometry

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Primary antibodies (CD45, CD14, CD68; all from Abcam, Cambridge, UK) were added to 100 μl samples of CBMCs, PLBMC, and MBMCs into 4 ml EDTA tubes. Tubes were vortexed for 5 s then left in the dark for 30 min followed by the addition of FITC-labeled secondary antibodies. Cells were then fixed in 500 μl of Optilyse C Solution (Beckman Coulter, High Wycombe, UK) added to all tubes which then were vortexed and left for 10 min. The final step was to add 500 μl of PBS to all tubes, followed by 100 μl of counting beads. The prepared tubes were left overnight at 4°C then analyzed using a Beckman Coulter FC500 flow cytometer. The bead count number was inserted into the protocol and the samples run on a low/medium speed. Gating criteria were set and data analyzed using WinMDI 2.8 software.
Maneta E., Warren A.Y., Hay D.P, & Khan R.N. (2015). Caspase-1-mediated cytokine release from gestational tissues, placental, and cord blood. Frontiers in Physiology, 6, 186.
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8

Competitive Binding of Monalizumab to HLA-E Tetramers

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Solution of biotinylated-HLA-E monomers (5nM solution, kindly provided by Novo Nordisk) was slowly mixed with PE-conjugated streptavidin (Southern Biotechnology) at a ratio 1/4 and then kept at 4°C until use. Competition of monalizumab and PE-conjugated tetramers was performed on peripheral blood from healthy donors (EFS). 100μL of whole blood was incubated for 1 h at 37°C with 50μL PBS containing PE-HLA-E tetramers (0.1μL) and monalizumab (ranging from 30μg/mL to 0,001μg/mL, 1/3 dilution) and then washed with PBS. Peripheral blood cells were then stained with anti-CD3-PercpCy5.5 (BD Biosciences), anti-CD45-APC (BD Biosciences), anti-CD3-PC7 (BD Biosciences) and anti-human IgG4 Fc-FITC (HP6025, Southern Biotechnology). Red blood cells were lyzed with Optilyse C solution (Beckman Coulter). Acquisition was performed on a FACSCanto II flow cytometer.
André P., Denis C., Soulas C., Bourbon-Caillet C., Lopez J., Arnoux T., Bléry M., Bonnafous C., Gauthier L., Morel A., Rossi B., Remark R., Breso V., Bonnet E., Habif G., Guia S., Lalanne A.I., Hoffmann C., Lantz O., Fayette J., Boyer-Chammard A., Zerbib R., Dodion P., Ghadially H., Jure-Kunkel M., Morel Y., Herbst R., Narni-Mancinelli E., Cohen R.B, & Vivier E. (2018). Anti-NKG2A mAb Is a Checkpoint Inhibitor that Promotes Anti-tumor Immunity by Unleashing Both T and NK Cells. Cell, 175(7), 1731-1743.e13.
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9

Multiparameter Flow Cytometry Immunophenotyping

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Blood collected into EDTA tubes was washed in PBS before staining with LiveDead (Thermo Fisher) according to the manufacturer’s instructions. Cells were incubated with mouse serum to saturate the Fc receptors, and were then incubated with the appropriate antibody cocktail. Red blood cells were lysed in Optilyse C Solution (Beckman Coulter), according to the manufacturer’s instructions. Cells were fixed in Cell Fix solution (BD), according to the manufacturer’s instructions. Data were acquired in an LSRFortessaX20 flow cytometer. The FCS3.0 files obtained were exported from BD FACSDiva software and imported into FlowJo v.10.5.2 (BD Biosciences). Automated compensation was calculated with FACSDiva software and single-stained compensation beads. This compensation matrix was analyzed in detail in FlowJo, by investigating the N-by-N view feature and the pairwise expression of all proteins stained in this study. Fluorescence minus one (FMO) experiments were run before this study, to facilitate optimization of the compensation matrix. We then adjusted the compensation matrix where necessary due to over- or under-compensation by the automatic algorithm.
Carvelli J., Demaria O., Vély F., Batista L., Benmansour N.C., Fares J., Carpentier S., Thibult M.L., Morel A., Remark R., André P., Represa A., Piperoglou C., Cordier P.Y., Le Dault E., Guervilly C., Simeone P., Gainnier M., Morel Y., Ebbo M., Schleinitz N, & Vivier E. (2020). Association of COVID-19 inflammation with activation of the C5a-C5aR1 axis. Nature, 588(7836), 146-150.
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