Rbs lysis buffer

Manufactured by BioLegend

RBS lysis buffer is a solution used for cell lysis and protein extraction. It is designed to efficiently disrupt cell membranes and release intracellular contents, including proteins, for subsequent analysis or purification.

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Lab products found in correlation

Collagenase type 1, by Merck Group (1 mentions) 40 μm strainer, by Thermo Fisher Scientific (1 mentions) Collagenase type 5, by Merck Group (1 mentions) Human fc block, by BD (1 mentions) Cd123 pe cy5, by BD (1 mentions) Aqua live dead stain, by Thermo Fisher Scientific (1 mentions) Cd19 pe, by BD (1 mentions) Hla dr pe cy7, by BioLegend (1 mentions) Dxp facscan, by Cytek (1 mentions) Cell strainer, by BD (1 mentions) Gentlemacs tissue dissociator, by Miltenyi Biotec (1 mentions)

Close spelling variants of this product

Lysis buffer

4 protocols using rbs lysis buffer

Extraction of Tumor Cells from Bone

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IC injected mice were euthanized according to IACUC guidelines. The method for tumor cell isolation from bone has been described in the RESULTs section. Briefly, muscle and connective tissue are removed from the femur and tibia. Bones were then ground with a mortar and pestle in 2 mg/ml Collagenase Type I (Sigma) dissolved in DMEM/F12. Ground bone pieces were cut into even smaller pieces with a scissor and transferred to bottles in a water bath for collagenase digestion at 37 °C. After a 25-min digestion, supernatant (1) was collected on ice and fresh collagenase digestion solution was added back for further digestion. After two more rounds of 25 min digestion, supernatant (2) and (3) were collected again on ice and the remaining bone pieces were washed with FACS buffer 3 times to collect all the released cells. Supernatant (1), (2), (3) and FACS buffer (0.5% BSA in PBS with 2 nM EDTA) collections were combined and filtered through 40 µm cell strainer to obtain a single cell suspension. Red blood cells were lysed using RBS lysis buffer (BioLegend) for 5 min on ice. For lung metastases, lungs were collected and cut into 1–2 mm³ pieces for digestion. Lung tissues were digested in 2 mg/ml collagenase solution for 45 min. Digested tissues were passed through a 40 µm cell strainer and subjected to RBC lysis as above.

Ren Q., Khoo W.H., Corr A.P., Phan T.G., Croucher P.I, & Stewart S.A. (2022). Gene expression predicts dormant metastatic breast cancer cell phenotype. Breast Cancer Research : BCR, 24, 10.

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Isolation of Pancreatic Macrophages

Cited 1 time

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Preparation of single-cell suspensions of pancreas was performed as previously described.⁴⁹ (link) Briefly, pancreata were minced using a pair of dissecting scissors, followed by digestion in 1 mg/mL collagenase type V (C9263; Sigma) in Hank’s balanced salt solution (SH3026801; Fisher) for 15 minutes at 37°C. Cells were washed in Hank’s balanced salt solution once, followed by incubation in RBS lysis buffer (420301; Biolegend, San Diego, CA) for 5 minutes. After passing through a 40-μm strainer (22363547; Fisher), single-cell suspensions were incubated with fluorescently conjugated antibodies (as shown in Table 1) diluted in fluorescence-activated cell sorter buffer (2% fetal bovine serum in Hank’s balanced salt solution). Cell analysis and sorting was performed using MoFlo Asterio sorter (Beckman Coulter, Indianapolis, IN). Macrophages (CD45⁺;CD11b⁺;F4/80⁺) were sorted and lysed in RLT plus buffer (QIA74136; Qiagen, Valencia, CA) for RNA isolation.

Wen H.J., Gao S., Wang Y., Ray M., Magnuson M.A., Wright C.V., Di Magliano M.P., Frankel T.L, & Crawford H.C. (2019). Myeloid Cell-Derived HB-EGF Drives Tissue Recovery After Pancreatitis. Cellular and Molecular Gastroenterology and Hepatology, 8(2), 173-192.

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Multiparametric Flow Cytometry Analysis

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After washing, treated cells and controls were incubated with human Fc-block (BD), and stained with the following antibodies as described in figure legends (at 1:100 dilution unless stated): IgE-FITC (eBiosciences clone: IgE21), FcɛRIα-APC (eBiosciences clone AER-37 at 1:50 dilution), anti-biotin AF-488 (eBiosciences clone: BK-1/39), anti-mouse-lambda-light-chain PE (BioLegend clone: RML-42), CD123 PE-Cy5 (BD clone: 9F5 at 1:20 dilution), HLA-DR PE-Cy7 (BioLegend clone L243 at 1:80 dilution), CD203c BV421 (BioLegend clone: NP4D6 at 1:50 dilution), CD19-PE (BD clone: HIB19 at 1:50), CD23-BV421 (BioLegend EBVCS-5 at 1:50 dilution) and Aqua Live Dead stain (Life Technologies). Stained cells were then lysed with RBS lysis buffer (BioLegend) for 5 min at room temperature, and washed with FACS buffer (PBS pH 7.4 supplemented with 10% FCS). Data were collected on a DxP FACSCAN from Cytek Development in Fremont, CA (10 colours with three lasers—488, 639, 407) using FlowJo CE and analysed using FlowJo (version 10).

Pennington L.F., Tarchevskaya S., Brigger D., Sathiyamoorthy K., Graham M.T., Nadeau K.C., Eggel A, & Jardetzky T.S. (2016). Structural basis of omalizumab therapy and omalizumab-mediated IgE exchange. Nature Communications, 7, 11610.

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Quantification of Lung Metastasis in Mice

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For lateral tail-vein injections, male NSG mice were injected with 5
× 10⁵ cells resuspended in 100 μl
PBS⁻ and lungs were analyzed for visible macroscopic
metastases formation 4–6 weeks after injection using a Leica MZ12
fluorescence dissection microscope. Lungs were fixed in 10% neutral buffered
formalin, embedded in paraffin, sectioned (5 μM), and stained with
H&E. Tumor cell extravasation was assessed on cryosections prepared 48
hours after tail-vein injections. Tdtom⁺ cells were scored as
extravasated if they were completely outside a capillary vessel and/or if
there was no overlap between their nuclei and CD31⁺ vessels. To
examine tdTom⁺ carcinoma cells in the lungs with flow cytometry,
lungs were minced and incubated in DMEM with 1.5 mg/ml collagenase A and
0.01% DNase for 30 minutes at 37°C and dissociated using a gentleMACS
tissue dissociator (Miltenyi Biotec). RBCs were lysed by resuspension in RBS
Lysis Buffer (BioLegend) and cells were passed through a cell strainer (BD
Biosciences) to obtain a single-cell suspension prior to flow cytometric
analysis. For metastasis experiments involving cells carrying
doxycycline-inducible constructs, expression was induced in
vitro for 5–7 days prior to tail-vein injection and mice
were maintained on a doxycycline-containing diet (625 mg/kg, Teklad/Envigo)
for the duration of the experiment.

Lambert A.W., Fiore C., Chutake Y., Verhaar E.R., Strasser P.C., Chen M.W., Farouq D., Das S., Li X., Eaton E.N., Zhang Y., Donaher J.L., Engstrom I., Reinhardt F., Yuan B., Gupta S., Wollison B., Eaton M., Bierie B., Carulli J., Olson E.R., Guenther M.G, & Weinberg R.A. (2022). ΔNp63/p73 Drive Metastatic Colonization by Controlling a Regenerative Epithelial Stem Cell Program in Quasi-mesenchymal Cancer Stem Cells. Developmental cell, 57(24), 2714-2730.e8.

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