Facs lysing solution

Manufactured by BD

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FACS lysing solution is a laboratory reagent used to prepare cell samples for flow cytometry analysis. It is designed to lyse red blood cells while preserving the integrity of the remaining cellular components, allowing for more accurate detection and analysis of specific cell populations.

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Lab products found in correlation

Facscanto 2, by BD (56 mentions) Facscalibur, by BD (51 mentions) Facsdiva software, by BD (51 mentions) Facscalibur flow cytometer, by BD (43 mentions) Brefeldin a, by Merck Group (39 mentions) Trucount tube, by BD (34 mentions) Facscanto 2 flow cytometer, by BD (33 mentions) Lsrfortessa, by BD (29 mentions) Lsrii flow cytometer, by BD (18 mentions) Cellquest software, by BD (17 mentions) Lsrfortessa flow cytometer, by BD (16 mentions) Cellfix, by BD (15 mentions) Facscanto flow cytometer, by BD (15 mentions) Perm wash buffer, by BD (13 mentions) Countbright absolute counting beads, by Thermo Fisher Scientific (13 mentions) Facscanto, by BD (12 mentions) Anti cd28, by BD (12 mentions) Ionomycin, by Merck Group (12 mentions) Phosphate buffered saline (pbs), by Thermo Fisher Scientific (11 mentions) Facsdiva, by BD (11 mentions)

Close spelling variants of this product

Erythrocyte FACS lysing solution BD fluorescence-activated cell sorter (FACS) lysing solution FACS Lysing solution 1X 500ul 1X BD FACS lysing solution BD FACS Lysing Lysing solution BD FACSTM

845 protocols using facs lysing solution

Immune Cell Response to LPS

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The following reagents were used: lipopolysaccharide from E. coli serotype 0111:B4 (LPS), fetal calf serum (FCS), dihydrorhodamine 123 (DHR), and phorbol 12-myristate 13-acetate (PMA) were purchased from Sigma Aldrich (Arklow, Ireland). Phycoerythrin labeled CD11b and BD FACS lysing solution were purchased from BD Biosciences (Oxford, UK). Alexa Fluor 647 antihuman TLR-4 was purchased from eBiosciences (Hatfield, UK). BD FACS lysing solution was purchased from BD Biosciences. Phosphate-buffered saline (PBS) was purchased from Oxoid, Thermo Fisher Scientific (Cambridge, UK). Dulbecco's modified Eagle's medium (DMEM), penicillin, streptomycin solution, and l-glutamate were purchased from GibcoBRL Life Technologies/ Invitrogen (Dublin, Ireland).

O'Hare F.M., Watson W., O'Neill A., Grant T., Onwuneme C., Donoghue V., Mooney E., Downey P., Murphy J., Twomey A, & Molloy E.J. (2015). Neutrophil and monocyte toll-like receptor 4, CD11b and reactive oxygen intermediates, and neuroimaging outcomes in preterm infants. Pediatric research, 78(1).

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Immune Cell Profiling of Candida Species

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C. albicans, C. glabrata, C. tropicalis, and C. parapsilosis strains were grown as previously described (strains and culture conditions). Aliquots were stained with FITC (fluorescein isothiocyanate), added at a concentration of 1 × 10⁶ cells per ml blood, and incubated at 37°C as indicated. To distinguish different immune cell populations, whole blood was stained with mouse anti-human CD3-PerCP (CD3-peridinin chlorophyll protein; clone SK7, T cells), CD19-PE (CD19-phycoerythrin; clone HIB19, B cells), CD45-PE-Cy7 (clone HI30, leukocytes), CD56-V450 (clone B159, NK cells), and CD66b-PE (clone G10F5; polymorphonuclear leukocyte [PMN]) obtained from BioLegend. Monocytes were stained with mouse anti-human CD14-PerCP (clone 47-3D6) from Abcam. Stained samples were treated with FACS lysing solution (BD), washed, and acquired immediately. For raw data analysis, FlowJo v10.0.8 software was used. The presence of activation markers was determined with mouse anti-human CD11b-V450 (clone ICRF44) from BD and CD16-BV510 (clone 3G8) and CD69-APC (CD69-allophycocyanin; clone FN50) from BioLegend. Stained samples were treated with FACS lysing solution (BD), washed, and acquired immediately. For raw data analysis, FlowJo v10.0.8 software was used.

Kämmer P., McNamara S., Wolf T., Conrad T., Allert S., Gerwien F., Hünniger K., Kurzai O., Guthke R., Hube B., Linde J, & Brunke S. (2020). Survival Strategies of Pathogenic Candida Species in Human Blood Show Independent and Specific Adaptations. mBio, 11(5), e02435-20.

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Cytokine Stimulation and Flow Cytometry Analysis

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Whole blood samples (540 μl) were supplemented with TNFα (#T6674, Sigma-Aldrich) to a final concentration of 10 ng/ml or left unstimulated. After incubation at 37°C for 20 min, the samples were transferred into FACS Lysing solution (diluted 1:10 in dH2O, #349202, BD Biosciences, Franklin Lakes, US-NJ), vortexed and placed on ice for 15 min before washing with PBS (277 rcf, 10 min, 4°C). After aspiration of the supernatant, the cells were resuspended in FACS Lysing solution (1:10, BD Biosciences) and the incubation and washing steps were repeated as previously. The cell pellets were resuspended in PBS and antibodies (all from BD Biosciences) were added in accordance with the dilutions recommended by the manufacturer; 1:20 for CD35 PE (#559872) and CD11b APC (#333143), and 1:40 for CD62L PE (#341012). One control per donor sample was left unlabeled. The samples were incubated in the dark at 4°C for 30 min and washed one more time in PBS before measurement in a flow cytometer (Accuri C6, BD Biosciences). The results were analyzed with the help of FlowJo 10 (BD Biosciences).

Sanchez Klose F.P., Björnsdottir H., Dahlstrand Rudin A., Persson T., Khamzeh A., Sundqvist M., Thorbert-Mros S., Dieckmann R., Christenson K, & Bylund J. (2021). A rare CTSC mutation in Papillon-Lefèvre Syndrome results in abolished serine protease activity and reduced NET formation but otherwise normal neutrophil function. PLoS ONE, 16(12), e0261724.

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Multiparameter Flow Cytometry for Eosinophil Identification

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Fluorochrome-labeled antibody mix was added to 50 μl of whole blood (about 2 × 10⁶ leukocytes) and incubated 30 min at room temperature in the dark [for example, anti-P2X4-FITC mAb27, anti-Siglec-8-PE (Nordic BioSite), anti-CD3-PerCP/Cy5.5 SK7 (BD), anti-CD20 PE L27 (BD), anti-CD14-APC cloneMφP9 (BD), anti-CD45-APC-Cy7 2D1 (BD)]. After two washes in PBS, stained samples were treated with FACS lysing solution for 10 min in the dark (Becton Dickinson), which lyses erythrocytes under gentle hypotonic conditions while preserving the leucocytes. Cells were then analyzed by FACS after a final wash in PBS. Eosinophils were identified as: CD45+ Siglec-8+ SSC high. Flow cytometry was performed with BD FACS Canto II (Figures 6, 7) and FACS data were analyzed using FlowJo, LLC.

Paalme V., Rump A., Mädo K., Teras M., Truumees B., Aitai H., Ratas K., Bourge M., Chiang C.S., Ghalali A., Tordjmann T., Teras J., Boudinot P., Kanellopoulos J.M, & Rüütel Boudinot S. (2019). Human Peripheral Blood Eosinophils Express High Levels of the Purinergic Receptor P2X4. Frontiers in Immunology, 10, 2074.

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Multiparametric Flow Cytometric Analysis

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Cell pellets were resuspended in PBS and stained with fluorescein isothiocyanate (FITC)-conjugated anti-CD4 (BD Biosciences) and allophycocyanin (APC)-conjugated anti-CD8 (BD Biosciences) antibodies; erythrocytes were lysed by the addition of FACS Lysing Solution (Becton Dickinson, San Diego, CA, USA). The cell suspensions were centrifuged, and the cell pellets were resuspended in PBS containing sodium azide and paraformaldehyde. Then, the cells were analysed with a FACSCalibur flow cytometer (Becton Dickinson) equipped with CellQuest software (Becton Dickinson). Cytospin samples were prepared using Auto Smear CF-12D (Sakura Co., Tokyo, Japan), and cellular infiltration in BAL fluid was assessed on Wright-Giemsa-stained slides (Wako Pure Chemical Industries, Ltd.).

Fujimoto Y., Hasegawa S., Matsushige T., Wakiguchi H., Nakamura T., Hasegawa H., Nakajima N., Ainai A., Oga A., Itoh H., Shirabe K., Toda S., Atsuta R., Morishima T, & Ohga S. (2017). Pulmonary inflammation and cytokine dynamics of bronchoalveolar lavage fluid from a mouse model of bronchial asthma during A(H1N1)pdm09 influenza infection. Scientific Reports, 7, 9128.

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Antigen Stimulation of Whole Blood Cells

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For antigen stimulation, we added 500 μL heparinized blood to a 5 mL polystyrene round-bottom tube (Corning, https://www.corning.com) containing a final concentration of 1 μL/mL anti-human CD28 and CD49d (Becton Dickinson, https://www.bd.com), along with B. pseudomallei, B. thailandensis, or BTCV antigens, all at a final concentration of 1 mg/mL. We used staphylococcal enterotoxin B (SEB) at a final concentration of 10 µg/mL as positive control and RPMI medium supplemented with 10% fetal bovine serum (FBS) (R10 medium) as negative control. We incubated the tubes at 37°C, 5% CO₂, 95% humidity. After 18 h, we added a final concentration of 10 μL/mL Brefeldin A (eBioscience, https://www.thermofisher.com) and incubated the assay for another 4–5 h under the same conditions. We then incubated samples with Live/Dead (LD) Fixable Near-IR Dead Cell Stain (Life Technologies, https://www.thermofisher.com) according to the manufacturer’s instructions, followed by a single 10-min incubation with FACS Lysing solution at room temperature for red cell removal (Becton Dickinson). We cryopreserved lysed blood cells in FBS with 10% dimethylsulfoxide (DMSO) (Sigma Aldrich, https://www.sigmaaldrich.com) and stored at −80°C until flow cytometry staining.

Rongkard P., Kronsteiner B., Hantrakun V., Jenjaroen K., Sumonwiriya M., Chaichana P., Chumseng S., Chantratita N., Wuthiekanun V., Fletcher H.A., Teparrukkul P., Limmathurotsakul D., Day N.P, & Dunachie S.J. (2020). Human Immune Responses to Melioidosis and Cross-Reactivity to Low-Virulence Burkholderia Species, Thailand. Emerging Infectious Diseases, 26(3), 463-471.

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Murine Models of Helminth Infection

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Female BALB/c and IL-4gfp 4get reporter mice on the BALB/c background were bred in-house and maintained under specific pathogen-free conditions at the University of Edinburgh. Mice were used at 6–12 weeks of age, and randomly assigned to experimental groups. The L. sigmodontis life cycle was maintained in gerbils (Meriones unguiculatus) using the mite vector Ornithonyssus bacoti [22 (link)]. Mice were infected s.c. on the upper back with 30 L. sigmodontis L3 larvae. Adult or larval parasites were recovered by lavage of the thoracic cavity. L. sigmodontis antigen (LsAg) was prepared by collecting the PBS-soluble fraction of homogenized adult male and female worms. To quantify blood microfilariae, 30 μL of tail blood was collected in FACS lysing solution (Becton-Dickinson), and microfilaria counted using a dark field optical microscopy (Axiovert 25, Zeiss). N. brasiliensis was maintained in Sprague-Dawley rats as previously described [23 (link)]. Mice were infected by s.c. injection with 200 N. brasiliensis L3 larvae. The parathymic, posterior, mediastinal and paravertebral LN, were taken as a source of thoracic LN (tLN) draining the pleural cavity (PleC). PleC cells were recovered by lavage. TLN cells were dissociated and washed in RPMI-1640 (invitrogen) supplemented with 0.5% mouse sera (Caltag-Medsystems), 100 U/ml penicillin, 100 μg/ml streptomycin and 2 mM L-glutamine.

Knipper J.A., Ivens A, & Taylor M.D. (2019). Helminth-induced Th2 cell dysfunction is distinct from exhaustion and is maintained in the absence of antigen. PLoS Neglected Tropical Diseases, 13(12), e0007908.

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Platelet Activation Surface Receptor Expression

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Expression of the platelet activation surface receptors glycoprotein IIb/IIIa, P-selectin, and CD63 was measured by flow cytometry. Healthy donor whole blood was collected in sodium citrate and treated with EPI (50 ng/ml) or control (HT buffer only) for either 3 min or 30 min of incubation. Blood was then diluted (1:50) with HT buffer, added to tubes with or without convulxin (5 ng/ml, Santa Cruz Biotechnology) and antibodies, and incubated at 37°C for 10 min. Allophycocyanin-conjugated anti-CD41 (clone HIP-8) was used to distinguish platelets, fluorescein isothiocyanate conjugated anti-CD62p (clone AK4) to distinguish platelets expressing surface P-selectin, anti-CD41/CD61 (clone PAC-1) to distinguish platelets with glycoprotein IIb/IIIa complex, and anti-CD63 (clone H5C6) to stain platelets expressing surface CD63. Staining with fluorophore and antibody isotype matched controls was performed to determine positive staining. All antibodies and immunoglobulin isotype controls were purchased from Biolegend. Stained samples were treated with FACS Lysing Solution at a ratio of 2:1 (Becton Dickinson) for 10 min at room temperature prior to storage at 4°C for no more than 24 h. Data were collected with a BD LSRII cytometer (Becton Dickinson) and analyzed using FlowJo (FlowJo, LLC).

Wang Z., Gergely J, & Tao T. (1992). Characterization of the Ca(2+)-triggered conformational transition in troponin C.. Proceedings of the National Academy of Sciences of the United States of America, 89(24), 11814-11817.

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Influenza Superinfection in Murine Filariasis

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Female BALB/c mice were purchased from Charles River and maintained under specific pathogen free conditions at the University of Edinburgh. Mice were used at 6–8 weeks of age. To maintain the L. sigmodontis lifecycle, L. sigmodontis infected jirds (Meriones unguiculatus) were used to infect haematophagous tick parasites (Ornithonyssus bacoti) in order to generate L3 stage larvae (23 ). Mice were infected s.c. on the upper back with 30 L. sigmodontis L3 stage larvae. L. sigmodontis larvae or adults were recovered from the pleural cavity by lavage and counted using a dissection microscope. To quantify blood Mf, 30 μL of tail blood was collected in FACS lysing solution (Becton-Dickinson) and the Mf counted using a haemocytometer. IAV infections were performed intranasally with 5x10³ PFU A/WSN/33, a mouse H1N1 influenza A strain (Dr D. Jackson, University of St Andrews, St Andrews, UK), either 12, 34 or 68 days (d) post L. sigmodontis or mock infection. A/WSN/33 was grown in MDCK cells as described previously (24 (link)). Mice were weighed daily and assessed for visual signs of clinical disease as described previously (25 (link), 26 (link)). Briefly, signs of infection were scored as follows, reduced mobility/activity (0-3), ruffled fur/piloerection (0-3), hunched posture (0-3) and increased or laboured breathing (0-3). The severity score presented is a sum of these criteria.

Hardisty G.R., Knipper J.A., Fulton A., Hopkins J., Dutia B.M, & Taylor M.D. (2022). Concurrent Infection With the Filarial Helminth Litomosoides sigmodontis Attenuates or Worsens Influenza A Virus Pathogenesis in a Stage-Dependent Manner. Frontiers in Immunology, 12, 819560.

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Evaluation of Cytotoxic Compounds

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The test compound DM (99% purity), low melting point agarose (LMPA), normal melting agarose (NMA), Na₂-EDTA, Trizma base, Ethidium bromide (EtBr), propiodium iodide (PI), RNAse, Rhodamine 123 (Rh123), electrophoresis reagents, sodium bicarbonate, phosphate buffered saline (PBS), and tris buffered saline were products of Sigma Aldrich (St. Louis, MO, USA). FACS Lysing Solution was a product of Becton Dickinson (San Jose, CA, USA). Bromophenol blue, Fetal bovine serum (FBS), and Colchicine were obtained from Hi-Media Pvt. Ltd. (Mumbai, India). Cyclophosphamide—CPA (Cyclocel 500) was obtained from Celon Laboratories (Andhra Pradesh, India). May—Gruenwald and Giemsa stain were procured from Merck (India).

Nazam N., Lone M.I., Hamid A., Qadah T., Banjar A., Alam Q., Saeed M, & Ahmad W. (2020). Dimethoate Induces DNA Damage and Mitochondrial Dysfunction Triggering Apoptosis in Rat Bone-Marrow and Peripheral Blood Cells. Toxics, 8(4), 80.

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