Washing buffer

Manufactured by BD

Sourced in United States

Washing buffer is a laboratory solution used to wash and rinse samples during various experimental procedures. It is designed to maintain the integrity and purity of the sample by removing unwanted materials, such as excess reagents, salts, or impurities. The core function of a washing buffer is to facilitate the cleaning and preparation of samples for further analysis or processing.

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Lab products found in correlation

Absolute methanol, by Merck Group (1 mentions) Streptomycin, by Merck Group (1 mentions) Penicillin, by Merck Group (1 mentions) Formaldehyde solution, by Merck Group (1 mentions) Concanavalin a (cona), by Merck Group (1 mentions) Sodium chloride (nacl), by Carl Roth (1 mentions) Kh2po4, by Carl Roth (1 mentions) Potassium chloride (kcl), by Carl Roth (1 mentions) Na2hpo4, by Carl Roth (1 mentions) L glutamine, by Harvard Bioscience (1 mentions) Facscanto 2 flow cytometer, by BD (1 mentions) Fix perm, by BD (1 mentions) Accuri c6 facs analyzer, by BD (1 mentions) Software package, by FlowJo (1 mentions) Hcd3 pe, by Thermo Fisher Scientific (1 mentions) Fcγr block, by Jackson ImmunoResearch (1 mentions) Hcd4 pe cy5, by BD (1 mentions) Facscalibur, by BD (1 mentions) Leukocyte activation cocktail, by BD (1 mentions)

Close spelling variants of this product

Permeabilization/washing buffer (2 citations)

4 protocols using washing buffer

T Cell Activation Assay Protocol

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Culture medium (CM) was prepared using RPMI 1640 supplemented with 100 U/mL of penicillin, 100 μg/mg streptomycin (both from Sigma, Steinheim, Germany), and 2 mM L-glutamine (Biochrom, Berlin, Germany). Concanavalin A (ConA) (Sigma) was diluted in CM to a concentration of 600 μg/mL and stored at −70°C. Antibodies were purchased from Becton Dickinson (BD, Heidelberg, Germany). Phosphate buffered saline (PBS) was made by dissolving 7.013 g NaCl, 0.2 g KCl, 1.513 g Na₂HPO4, and 0.2 g KH₂PO4 (all purchased from Roth) in 1 liter distilled water and by adjusting the pH to 7.4. Red blood cell (RBC) lysing solution was purchased from Becton Dickinson and diluted 1 : 10 in distilled water before use. Washing buffer was obtained from BD Biosciences (San Diego, USA). Formaldehyde solution and absolute methanol was purchased from Merck (Darmstadt, Germany).

Dieterlen M.T., Bittner H.B., Tarnok A., Garbade J., Dhein S., Mohr F.W, & Barten M.J. (2014). Flow Cytometric Evaluation of T Cell Activation Markers after Cardiopulmonary Bypass. Surgery Research and Practice, 2014, 801643.

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Platelet Flow Cytometry Protocol

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Flow cytometric analyses were done using washed platelets obtained from EDTA whole blood. To analyze intrinsic proteins washed platelets were fixed and permeabilized in Fix/Perm (BD Biosciences) solution for 20min at RT. Afterwards, platelets were washed with washing buffer (10x Perm/Wash Buffer; BD Biosciences) and centrifuged for 10min at 500g. All washing steps were performed with a washing buffer from BD and followed by 500g centrifugation for 10min at RT. To block nonspecific binding, blocking buffer (PBS+5% FBS) was added to the platelets and incubated for 30min at RT. Supernatant was removed and the primary antibody was added for 30min at RT. After removing the supernatant, secondary antibodies (1:1000 dilution) were diluted in PBS + 2% FBS and incubated for 30min at RT. The platelets surface marker CD42a- or b- PE was added and incubated for 30min at RT. Detailed antibody list is presented in S1 Table. For each sample, 10’000 platelets were identified as CD42a- or CD42b-positive events. All samples were acquired using a FACS Canto II flow cytometer (Becton Dickinson, Rotkreuz, Switzerland). Data was analyzed with FlowJo software (version 10.0).

Goelz N., Eekels J.J., Pantic M., Kamber C.T., Speer O., Franzoso F.D, & Schmugge M. (2021). Platelets express adaptor proteins of the extrinsic apoptosis pathway and can activate caspase-8. PLoS ONE, 16(1), e0244848.

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Assessing Humanized Mouse Engraftment by Flow Cytometry

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Flow cytometry was used to assess human cell engraftment levels in humanized SIVsm infected and uninfected mice. Peripheral blood was collected in heparinized capillary tubes by tail vein puncture bimonthly. 5 ul of FcγR-block (Jackson ImmunoResearch Laboratories, Inc. West Grove, PA) was added to the blood cell pellet and then stained with fluorophore conjugated hCD45-FITC (eBioscience), hCD3-PE (eBioscience) and hCD4-PE/CY5 (BD Pharmingen, San Jose, CA) for 30 min. Erythrocytes were lysed using a RBC lysing kit (BD sciences). After lysing, stained cells were washed twice with washing buffer (BD Sciences) and fixed in 1% paraformaldehyde. Stained cells were analyzed using BD Accuri C6 FACS analyzer. In order to measure CD4⁺ T cell depletion in SIV infected mice, CD3⁺ T cell levels were calculated as a ratio of the entire CD45⁺ (lymphocyte common antigen marker) population. The CD4⁺ T cell level was then determined based on the ratio of the entire CD3⁺ T cell population‥ Engraftment levels of CD45, CD3 and CD4 were measured before infection as a control. CD4⁺ T cell decline was assessed using a two-tailed Student’s t-test (<0.05) to compare the two groups of mice, infected and uninfected. All flow data was analyzed using the FlowJo software package (FlowJo LLC, Ashland, Oregon).

Schmitt K., Kumar D.M., Curlin J., Remling-Mulder L., Stenglein M., O’Connor S., Marx P, & Akkina R. (2017). Modeling the evolution of SIV sooty mangabey progenitor virus towards HIV-2 using humanized mice. Virology, 510, 175-184.

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Quantification of T Cell Subsets

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Flow cytometry was performed to detect the quantitative proportion of CD4⁺ Tim‐3⁺ T cells and CD4⁺ IL‐17⁺ T cells. Murine spleens were collected, and lymphocytes were resuspended with 100 μL of RPMI 1640 medium and incubated with 2.5 μg·mL⁻¹ PMA, 1.0 μg·mL⁻¹ Ionomycin, and 10 μg·mL⁻¹ BFA for 4 h in a humidified incubator with 37 °C and 5% carbon dioxide. In addition, 200 μL aliquots of human peripheral blood samples from each group were diluted with 800 μL of RPMI 1640 medium and incubated with 2 μL Leukocyte Activation Cocktail (BD Bioscience) in a humidified incubator with 37 °C and 5% carbon dioxide for 4 h. After incubating the murine samples with Tim‐3 APC and CD4 FITC mouse antibodies, and human samples with Tim‐3 BB515 and CD4 APC human antibodies for 20 min in the dark at room temperature, the cells were washed twice with PBS buffer, and incubated with 250 μL fixed membrane breaker (BD Bioscience) along with mice samples for 20 min. They were then washed twice with washing buffer (BD Bioscience), incubated with IL‐17 PE mouse antibody for 20 min, and washed twice with PBS buffer. Subsequently, the cells were analyzed using a FACSort cell analyzer (FACSCalibur; Becton‐Dickinson, San Jose, CA, USA) according to the manufacturer's protocol.

Wu H., Tang S., Zhou M., Xue J., Yu Z, & Zhu J. (2021). Tim‐3 suppresses autoimmune hepatitis via the p38/MKP‐1 pathway in Th17 cells. FEBS Open Bio, 11(5), 1406-1416.

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