Rbc lysis buffer solution

Manufactured by Thermo Fisher Scientific

Sourced in United States

RBC Lysis Buffer Solution is a laboratory reagent designed to selectively lyse red blood cells (RBCs) while preserving the integrity of other cell types, such as leukocytes. The solution contains a proprietary combination of compounds that effectively rupture RBC membranes, allowing for the separation and isolation of non-RBC components from whole blood samples.

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Lab products found in correlation

2 mercaptoethanol, by Thermo Fisher Scientific (2 mentions) Easysep mouse monocyte enrichment kit, by STEMCELL (1 mentions) Liberase dl, by Roche (1 mentions) Dnase, by Roche (1 mentions) Mcp 1, by R&D Systems (1 mentions) Rpmi medium, by Thermo Fisher Scientific (1 mentions) Dulbecco phosphate buffered saline (dpbs), by Corning (1 mentions) Carboxyfluorescein succinimidyl ester (cfse), by Thermo Fisher Scientific (1 mentions) Cd4 e450, by Thermo Fisher Scientific (1 mentions) Propidium iodide staining solution, by Thermo Fisher Scientific (1 mentions) Cd16 32 fc block, by BioLegend (1 mentions) Cd8α apc, by Thermo Fisher Scientific (1 mentions) Siinfekl, by MBL Life Science (1 mentions) Collagenase, by Merck Group (1 mentions) Cell lysis buffer, by Cell Signaling Technology (1 mentions) Qiazol lysis reagent, by Qiagen (1 mentions) Anti cd44 buv395, by BD (1 mentions) Anti pd l1 bv421, by BioLegend (1 mentions) Anti cd25 pe, by BioLegend (1 mentions) Anti cd8 apc, by BD (1 mentions)

Close spelling variants of this product

RBC lysis buffer RBC lysis solution RBC lysis Lysis buffer Lysis solution Buffer solutions

9 protocols using rbc lysis buffer solution

Monocyte Migration Assay Protocol

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Spleens were mechanically dissociated and the tissue digested with Liberase DL and DNase (Roche Diagnostics) [12 (link)]. Red cells were lysed using 1X RBC Lysis Buffer Solution (eBioscience, San Diego, CA). Monocytes were enriched by negative selection using the Easy Sep Mouse Monocyte Enrichment Kit (Stem Cell Technologies, Cambridge, MA). Monocytes (1×10⁶) were placed within 3 μm pore cell culture inserts (Corning, Corning, NY). Wells below the inserts were supplemented with or without 10 ng/ml MCP-1 (R&D Systems, Minneapolis, MN). Cells that migrated through the pores were stained using crystal violet and counted.

Trial J., Heredia C.P., Taffet G.E., Entman M.L, & Cieslik K.A. (2017). Dissecting the Role of Myeloid and Mesenchymal Fibroblasts in Age-dependent Cardiac Fibrosis. Basic research in cardiology, 112(4), 34.

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Generation of GM-CSF and FLT3L Dendritic Cells

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BMDCs were prepared from 6–10 week-old C57BL/6J female femurs and tibias as previously described.²⁴ (link) Briefly, bone ends were cut off and bone marrow was flushed with RPMI medium (GIBCO, 21875–034) containing 5% FCS and 50 mM of 2-mercaptoethanol (GIBCO, 31350–010). Red blood cells were removed by 1 min exposure to 1xRBC lysis buffer solution (eBioscience, 00–4333–57). For GM-DCs, 3 × 10⁶ cells were seeded onto 6-well plates in 5 mL medium containing 0.8 % supernatant of the J558L GM-CSF producing cell line. Medium was changed at day 2.5 and GM-DCs were ready to use at day 5. For FL-DCs, 8 × 10⁶ cells were seeded onto 6-well plates in 4 mL medium containing 3% supernatant of the B16 FLT3 Ligand producing cell line. FL-DCs were ready to use at day 8.5–9 without any change of medium.

Zhao Y., Hanniffy S., Arce-Gorvel V., Conde-Alvarez R., Oh S., Moriyón I., Mémet S, & Gorvel J.P. (2017). Immunomodulatory properties of Brucella melitensis lipopolysaccharide determinants on mouse dendritic cells in vitro and in vivo. Virulence, 9(1), 465-479.

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Isolation and Stimulation of Murine Splenocytes

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Spleen and draining lymph nodes (LNs) were harvested and prepared for cell culture by processing through a 100-μm cell strainer with RPMI 1640 media supplemented with 10% heat-inactivated fetal bovine serum (Hyclone, Logan, UT), 100-μg/mL streptomycin, 100 U/mL penicillin, 50-μM 2-mercaptoethanol, 25-mM HEPES, and 2-mM l-glutamine, 1-mM sodium pyruvate, and 1× nonessential amino acids (Mediatech, Manassas, VA). Red blood cells within splenocyte preparation were lysed using RBC Lysis Buffer Solution (eBioscience, San Diego, CA). Cells were plated at a concentration of 2 × 10⁵ cells per 200 μL final volume in round-bottom 96-well culture plates. Cells were stimulated with either 1-μg/mL anti-CD3ε (2C11; R&D, Minneapolis, MN) or 2-μg/mL Dby peptide (AnaSpec, Fremont, CA). Culture supernatants were harvested on day 4 of culture, and IL-17A and IFNγ concentrations were measured by ELISA (R&D, Minneapolis, MN) according to manufacturer's instructions.

Julliard W., Fechner J.H., Owens L., O'Driscoll C.A., Zhou L., Sullivan J.A., Frydrych L., Mueller A, & Mezrich J.D. (2017). Modeling the Effect of the Aryl Hydrocarbon Receptor on Transplant Immunity. Transplantation Direct, 3(5), e157.

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In vivo SPIO NWs Trafficking in Mice

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Peripheral blood was collected from BALB/c mice by cardiac puncture using heparin as anticoagulant. Blood was centrifuged at 2000g for 5 min at room temperature, the supernatant was discarded and red blood cells were lysed with RBC Lysis Buffer Solution (eBioscience, San Diego, USA). Leukocytes were washed 2 times with 1 × DPBS (Corning, Manassas, USA). Leukocytes (1 × 10⁶ cells/mL) were labeled with 10 μM carboxyfluorescein succinimidyl ester CFSE (eBioscience) at room temperature for 8 min. Reaction was blocked with 2% fetal calf serum, and the cells were washed in 1 × DPBS 2 times. Resuspended leukocytes were incubated with 0.1 mg/mL SPIO NWs (preincubated with mouse serum as described above) at 37 °C with gentle shaking for 1 h. SPIO NWs labeled leukocytes were isolated with MiniMACS column; both magnetic and non magnetic fraction were collected. Mice were injected with 0.5–1 × 10⁶ leukocytes in 100 μL PBS via intravenous injection. Blood was collected from the periorbital plexus at 1 min, 10 min, 60 min, 240 and 1440 min postinjection and CFSE positive cells were counted with hemocytometer.

Inturi S., Wang G., Chen F., Banda N.K., Holers V.M., Wu L., Moghimi S.M, & Simberg D. (2015). Modulatory Role of Surface Coating of Superparamagnetic Iron Oxide Nanoworms in Complement Opsonization and Leukocyte Uptake. ACS nano, 9(11), 10758-10768.

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Isolation of Bone Marrow Cell Fractions

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Bone cells were isolated for mRNA analysis as described.^{(9 (link))} Briefly, flushed bone marrow cells were pooled and treated with red blood cell (RBC) lysis buffer solution (eBioscience, San Diego, CA, USA) for 5 min at room temperature. Following a brief centrifugation and resuspension in FACS buffer, the resulting bone marrow mononuclear cells (BMMNCs) were further processed using magnetic activated cell sorting (MACS; autoMACS-Pro magnetic cell sorter; Miltenyi Biotec, Inc., San Diego, CA, USA) to obtain highly enriched fractions of myeloid cells, osteoblasts, and osteocytes, as described.^{(9 (link))}

Chandra A., Lagnado A.B., Farr J.N., Monroe D.G., Park S., Hachfeld C., Tchkonia T., Kirkland J.L., Khosla S., Passos J.F, & Pignolo R.J. (2020). Targeted Reduction of Senescent Cell Burden Alleviates Focal Radiotherapy-Related Bone Loss. Journal of bone and mineral research : the official journal of the American Society for Bone and Mineral Research, 35(6), 1119-1131.

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Quantification of Antigen-Specific T Cells

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At different time points after the immunization, blood was collected via mouse tail vein, and the red blood cells were lysed using a RBC lysis buffer solution (eBioscience, San Diego, CA). The monocytes were incubated with Fc block (CD16/32, BioLegend, diluted in FACS buffer at 1:9) at 4 °C for 15 min. The monocytes were then incubated for 30 min at room temperature with PE conjugates of MHC tetramers specific for either CD4 T cells recognizing the OVA epitope ISQAVHAAHAEINEAGR (MBL International, Woburn, MA), CD 8 T cells recognizing the OVA epitope SIINFEKL (MBL International), CD 8 T cells recognizing the gp 100 epitope EGSRNQDWL (MBL International), or CD 8 T cells recognizing the TRP2 epitope SVYDFFVWL (MBL International). Subsequently, the cells were incubated for 10 min at room temperature with a propidium iodide staining solution (eBioscience, diluted in FACS buffer 1:25), CD4-e450 (eBioscience, diluted in FACS buffer 1:200), and CD8α-APC (eBioscience, diluted in FACS buffer 1:200). The cells were washed with FACS buffer, and data were collected on a BD LSR II.

Oberli M.A., Reichmuth A.M., Dorkin J.R., Mitchell M.J., Fenton O.S., Jaklenec A., Anderson D.G., Langer R, & Blankschtein D. (2016). Lipid Nanoparticle Assisted mRNA Delivery for Potent Cancer Immunotherapy. Nano letters, 17(3), 1326-1335.

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Adipose Tissue Fractionation and Analysis

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Epididymal white adipose tissue (eWAT) was extracted from 19 LFD-fed WT mice, 19 HFD-fed WT mice, 7 LFD-fed LysMcre(+/−).Atg16l1^flox/flox mice, 7 HFD-fed LysMcre(+/−).Atg16l1^flox/flox mice, 5 LFD-fed LysMcre(−/−).Atg16l1^flox/flox mice, and 6 HFD-fed LysMcre(−/−).Atg16l1^flox/flox mice. eWAT was washed once in FACS Buffer (dPBS buffer containing 0.2% BSA (Sigma, A2153) 1% EDTA) and centrifuged at 500g x 5 min to remove peripheral red blood cells. Tissue was digested in collagenase (Sigma, C6885) at 37°C for 30 min – 1 h to obtain a single cell suspension. Cells were filtered (100 μm) and centrifuged (500g x 5 min at 4°C) to separate the floating adipocyte fraction from the stromal vascular fraction (SVF). The floating adipocyte fraction from 8 LFD-fed WT mice and 8 HFD-fed WT mice were removed and placed into QIAzol lysis reagent (QIAGEN, 79306) for RNA extraction or 1X cell lysis buffer (Cell Signaling, 9803) for protein extraction. SVFs were incubated with RBC lysis buffer solution (eBioscience, 00-4333-57) for 3 min at room temperature and then centrifuged at 500g x 5 min at 4°C.

Litwinoff E.M., Gold M.Y., Singh K., Hu J., Li H., Cadwell K, & Schmidt A.M. (2017). Myeloid ATG16L1 does not affect adipose tissue inflammation or body mass in mice fed high fat diet. Obesity research & clinical practice, 12(2), 174-186.

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In Vitro Generation of OVA-Specific Memory CD8+ T Cells

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In Vitro Memory Differentiation Memory OT-I T cells were generated as described previously (Balmer et al., 2016; van der Windt et al., 2013) . Briefly, the lymph nodes from MHC class I-restricted OVA-specific T cell receptor (OT-I) transgenic mice and the spleen of C57BL/6 mice were aseptically removed and incubated in liberase TL (Roche) for 30 min. After mashing through a 70 mm cell strainer (BD Biosciences), red blood cells were lysed with RBC Lysis Buffer Solution (eBioscience). The isolated cell suspensions were washed in RPMI medium (RPMI 1640 containing 10% FCS, 100 U/mL penicillin, 100 mg streptomycin, 0.29 mg/mL L-glutamine, 50 mM 2-Mercaptoethanol (Life Technologies)) and re-suspended to 10 6 cells/mL. The splenocytes and OT-I cells were pooled in a ratio of 1:1 and activated with OVA peptide (Eurogentec) at 10 À9 M at 37 C for 3 days. The cells were then washed and re-suspended to 2 3 10 6 cells/mL and cultured in the presence of IL-15 (10 ng/mL) at 37 C for another 3 days to generate OVA-specific memory CD8 + T cells. Phenotyping was performed using BUV395-anti CD44 (BD), APC-anti CD8, BV421-anti KLRG1, PE-anti-CD62L, APC-Cy7-anti CD43, BV421-anti PDL1, PE-anti-CD25, BV510-anti CD27 (all Biolegend) and PE-Cy5-anti CD127 (eBioscience).

Balmer M.L., Ma E.H., Thompson A.J., Epple R., Unterstab G., Lötscher J., Dehio P., Schürch C.M., Warncke J.D., Perrin G., Woischnig A.K., Grählert J., Löliger J., Assmann N., Bantug G.R., Schären O.P., Khanna N., Egli A., Bubendorf L., Rentsch K., Hapfelmeier S., Jones R.G, & Hess C. (2020). Memory CD8⁺ T Cells Balance Pro- and Anti-inflammatory Activity by Reprogramming Cellular Acetate Handling at Sites of Infection. Cell metabolism, 32(3).

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Isolation and Analysis of Rat Peripheral Blood Leukocytes

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Blood samples were collected 1 week before rats were sacrificed by punctuation of the tail vein. Approximately 500 μl of peripheral blood were collected in BD Microtainer Plastic Capillary Blood Cellectors with Dipotassium EDTA. Erythrocytes were lysed with 1 ml of RBC Lysis buffer solution (eBioscience, Cat.-No. 00-4333-57) for 5 min at 37°C. The lysis reaction was neutralized with 10 ml of saline. Peripheral blood leukocytes were collected by centrifugation at 500 g for 5 min. The resulting pellets were resuspended in 400 μl of cold flow cytometry staining buffer solution and processes for flow cytometry. Samples were incubated for 30 min on ice with TruStain fcX to block Fc receptors and subsequently incubated with a cocktail of fluorophore-labeled monoclonal antibodies for 1 h at 4°C protected from light. Following a wash with ice-cold saline, samples were finally resuspended in 400 μl of cold Flow Cytometry Staining Buffer Solution. Different antibodies were used to define the cell populations as indicated in Table 1. Data were acquired on an LSRII flow cytometer (BD Bioscience) and analyzed with FlowJo software, version 7.6.3.

Schreckenberg R., Wolf A., Troidl C., Simsekyilmaz S, & Schlüter K.D. (2021). Pro-inflammatory Vascular Stress in Spontaneously Hypertensive Rats Associated With High Physical Activity Cannot Be Attenuated by Aldosterone Blockade. Frontiers in Cardiovascular Medicine, 8, 699283.

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