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Nh4cl solution

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NH4Cl solution is a laboratory reagent that provides a source of ammonium ions (NH4+) in aqueous solutions. It is a colorless, odorless liquid that is commonly used in various scientific applications.

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Lab products found in correlation

Penicillin, by Thermo Fisher Scientific (2 mentions) Streptomycin, by Thermo Fisher Scientific (2 mentions) Mouse cell depletion kit, by Miltenyi Biotec (1 mentions) Penicillin streptomycin, by Thermo Fisher Scientific (1 mentions) Facsaria 2, by BD (1 mentions) Dispase, by STEMCELL (1 mentions) Fetal calf serum (fcs), by STEMCELL (1 mentions)

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NH4Cl

4 protocols using nh4cl solution

1

Murine Hematopoietic Cell Isolation and Analysis

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Murine BM cells were flushed out of femurs, tibiae, hip bone and humeruses using a syringe filled with DPBS and filtered through a 70 μm-cell strainer. After red blood cell lysis using NH4Cl solution (STEMCELL Technologies), mononuclear cells were phenotyped using different antibody cocktails from Biolegend, e-Bioscience or Beckton Dickinson. Flow cytometry analysis was performed with a BD FACS LSRIITM flow cytometer (BD Biosciences) and cell sorting with a FACS Influx cell sorter (Becton Dickinson). Data were analyzed with FlowJo software. Antibodies and gating strategies for hematopoietic subset analysis and sorting are described in the Online Supplementary Methods. For RT-PCR and Ape1 endonuclease activity experiments, aliquots of 50,000 myeloid progenitor cells were sorted in PBS whereas aliquots of 10,000 HSC and multi-potent progenitors (MPP) were sorted in PBS/1%BSA.
Siberchicot C., Gault N., Déchamps N., Barroca V., Aguzzi A., Roméo P.H., Radicella J.P., Bravard A, & Bernardino-Sgherri J. (2020). Prion protein deficiency impairs hematopoietic stem cell determination and sensitizes myeloid progenitors to irradiation. Haematologica, 105(5), 1216-1222.
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2

Isolation of Mouse Bone Marrow Cells

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Mouse bone marrow cells were isolated from the femoral bones of BALB/c mice as described previously with slight modifications [26 (link)]. In brief, femoral bones were removed from CO2-euthanized mice under sterile conditions and immersed in ice-cold Hank's balanced salt solution (HBSS) containing 100 U/mL penicillin and 100 μg/mL streptomycin (all from Thermo Fisher Scientific). Both epiphyses of the femurs were removed with sterile scissors, and bone marrow cells were collected by strongly flushing the diaphysis with ice-cold HBSS using a 1 mL syringe. RBCs were lysed using a 0.9% (w/v) NH4Cl solution (Stemcell Technologies, Cambridge, MA, USA). After washing with HBSS, bone marrow cells were resuspended in RPMI-1640 media containing 2% FBS, 100 U/mL penicillin, and 100 μg/mL streptomycin. Viable cells were counted using the trypan blue exclusion method.
Lin S.H., Li M.H., Chuang K.A., Lin N.H., Chang C.H., Wu H.C., Chao Y.H., Lin C.C., Pan I.H., Perng M.D, & Wen S.F. (2020). Chlorella sorokiniana Extract Prevents Cisplatin-Induced Myelotoxicity In Vitro and In Vivo. Oxidative Medicine and Cellular Longevity, 2020, 7353618.
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3

Quantifying Pinocytosis in Leukocytes

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Pinocytosis was assessed via Lucifer Yellow (LY, Biotium) uptake, as described previously [22 (link)], with some modifications. For in vivo samples, a NH4Cl solution (StemCell Technologies) was used to lyse erythrocytes from whole blood, yielding blood leukocytes. Airway cells were obtained as described above. Blood leukocytes and airway cells were then stained with probes for viability (Live/Dead) and antibody markers for PMNs (CD66b) and exocytosis (CD63), as described above. Cells were then incubated with LY diluted at 1 mg/mL in RPMI for 30 mins at 37ºC. For in vitro samples, PMNs were transmigrated to CFASN containing LY (1 mg/mL) for 2, 10, and 18 hours, after which the cells were washed with PBS-EDTA and stained with viability probes and antibodies, as above. Cells were then washed with RPMI and pinocytosis was measured by flow cytometry.
Forrest O.A., Ingersoll S.A., Preininger M.K., Laval J., Limoli D.H., Brown M.R., Lee F.E., Bedi B., Sadikot R.T., Goldberg J.B., Tangpricha V., Gaggar A, & Tirouvanziam R. (2018). PATHOLOGICAL CONDITIONING OF HUMAN NEUTROPHILS RECRUITED TO THE AIRWAY MILIEU IN CYSTIC FIBROSIS. Journal of leukocyte biology, 104(4), 665-675.
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4

Tumor Cell Isolation and Sorting

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A two-step digestion protocol was used to improve dissociation and isolation of tumor cells. First, excised xenograft tumors were minced and dissociated in the 37°C shaker for 2 hours with medium containing 10% Fetal Calf Serum, 0.5 U/ml dispase (#07913, STEMCELL Tech.), 5mg/ml Collagenase Type IV (CLS-4, Worthington Biochem.), and 100 U/ml Penicillin Streptomycin (15-140-122, Gibco) in DMEM (11965-084, ThermoScientific). The digested tumor tissues were pelleted and washed once in PBS before they were resuspended in 0.25% trypsin and briefly digested at room temperature for <5 minutes by repeated gentle pipetting. The dissociated cells were then collected by filtering the digested tissues through a 70-μm cell strainer. Red blood cells were removed using an NH4Cl Solution (07800, STEMCELL Tech.). Host mouse cells were depleted using the Mouse Cell Depletion Kit (130-104-694, Miltenyi Biotec.) EGFP+ (hypoxic) and EGFP− (non-hypoxic) tumor cells were sorted using BD FACSAria™ II.
Kim H., Lin Q, & Yun Z. (2018). The hypoxic tumor microenvironment in vivo selects the tumor cells with increased survival against genotoxic stresses. Cancer letters, 431, 142-149.
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