Выделение и характеристика химерных человека Fc-экспрессии белков с использованием белок мембраны адсорберов и рационализировать рабочий процесс


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Burdick, M. M., Reynolds, N. M., Martin, E. W., Hawes, J. V., Carlson, G. E., Cuckler, C. M., Bates, M. C., Barthel, S. R., Dimitroff, C. J. Isolation and Characterization Of Chimeric Human Fc-expressing Proteins Using Protein A Membrane Adsorbers And A Streamlined Workflow. J. Vis. Exp. (83), e51023, doi:10.3791/51023 (2014).

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Name Company Catalog Number Comments
Protein A membrane adsorber (Sartobind Protein A 2 ml)  Sartorius-Stedim 93PRAP06HB-12--A
Amicon Ultra centrifugal filter, 10 kDa, 4 ml Millipore UFC801008 Use 15 ml volume if needed
Sterile vacuum filter unit, 0.22 µm, 500 ml Millipore SCGPU05RE
10 ml Syringes with Luer Lock tips BD 301604
30 ml Syringes with Luer Lock tips BD 309650
Tygon lab tubing (1/16 in x 1/8 in x 50 ft) Cole Parmer WU-95903-16
Slide-A-Lyzer MINI Dialysis Devices (10K MWCO) Thermo Scientific 69576
Snap i.d. antibody collection tray EMD Millipore WBAVDABTR
Snap i.d. single well blot holder EMD Millipore WBAVDBH01
Elution buffer Thermo Scientific 21004
Tris, ultra pure Fisher Scientific 819623
Sodium chloride Fisher Scientific 7647-14-5
Tween 20 Fisher Scientific BP337-100
DPBS Thermo Scientific SH30028.02
DPBS with Ca2+/Mg2+ Life Technologies 14080-055
BSA Sigma A9647
Human Fc Bethyl Research Laboratories P80-104
Anti-human Fc-APC Jackson Immunoresearch 109136170
Anti-human Fc-AP Bio-Rad 170-5018
Alkaline phosphatase substrate Promega S3841
Flow cytometry tubes (5 ml polystyrene or polypropylene) BD 352054
Syringe pump Harvard Apparatus 55-2226 A peristaltic pump is also acceptable
Protein gel electrophoresis system Bio-Rad Laboratories 552BR094876 Any gel electrophoresis system is acceptable, but mini gel systems run fastest
Semidry blotter Bio-Rad Laboratories 690BR006163 Any transfer system is acceptable, but discontinuos transfer systems perform electrophoretic transfer fastest
SNAP i.d. vacuum-assisted protein detection system EMD Millipore WBAVDBASE An upgraded model has replaced this specific instrument, but either should work just as well



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