Concentration of Phage Lysate using Vivaflow Tangential/Crossflow Filtration Cassette v1 [dataset]

Sarah Giuliani
2016 protocols.io  
Vivaflow 50 cassette, (100,000 MWCO, PES),which includes size 16 tubing for inlet and outlet sample flow, and interconnector tub VF05P4 Pressure Indicator, must order separately for Vivaflow 50 VFA020 Masterflex console drive pumps (Cole-Parmer): 1 per phage sample Masterflex Easy Load II Model 7702-50 pump head (tubing size LS 13.14.16.25): 1 per pump -ours have capacity to load two separa Masterflex tubing of size 25, 2 ft length (cat# 96410-25): (cleaned, acid washed, autoclaved -wrapped in
more » ... oil), 1 per Vivaflow 2 Filtrate collection bottles or flasks (acid wash container if keeping phage filtrate samples): 1 per phage sample 250 ml polycarbonate bottles: 3 per 2 Vivaflow 50 cassettes (for each phage) -for sample recovery and rinsing steps 2 sterile 50 ml conical tubes Parafilm 3 L of Nanopure water, 0.2 um filtered into acid washed bottles and autoclave sterilized: use to pre-rinse new Vivaflow cassette 600 ml ASW salts media 1.5 ml Eppendorf tubes (autoclaved) -for SYBR slide sample collection BEFORE STARTING 1 1 Coleman Lab Purpose Purpose : To concentrate harvested phage lysate up to 20X -40X. The Vivaflow device uses a peristaltic pump setup to recirculate the input sample over a 100,000 MWCO PES membrane, which allows the media filtrate to be diverted and the phage to remain in the starting sample. MODIFICAT ION NOT ES: MODIFICAT ION NOT ES: To ensure full phage recovery from large sample starting volumes of up to 4-8 L of lysate, some steps were added to the manufacturer's instructions based on empirical trials of filtering capacity. The first trial was done following the Sartorius user manual steps explicitly for 8 L filtered on a Vivaflow 200 device, but resulted in about 25% phage recovery. The second trial incorporated modifications which enhanced recovery to nearly 100% from 4 L of lysate using a Vivaflow 200 device and 2 L of lysate using a Vivaflow 50 device, and is described in this protocol for the Vivaflow 50. This may not be an issue for sample volumes of less than 2 L being concentrated on the same device (Vivaflow 50), but this was not verified.
doi:10.17504/protocols.io.fhdbj26 fatcat:hxrusoghsbamxjivcsxcalv3hm