May 13, 2023

Public workspaceNCEM Drop - Cell Pellet Dounce Homogenisation (TM-014) V.2

DOI
dx.doi.org/10.17504/protocols.io.bp2l69j4dlqe/v2
  • 1CSIRO ACDP
sandra.crameri
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DOI: dx.doi.org/10.17504/protocols.io.bp2l69j4dlqe/v2
Protocol Citation: sandra.crameri 2023. NCEM Drop - Cell Pellet Dounce Homogenisation (TM-014). protocols.io https://dx.doi.org/10.17504/protocols.io.bp2l69j4dlqe/v2Version created by sandra.crameri
License: This is an open access protocol distributed under the terms of the Creative Commons Attribution License,  which permits unrestricted use, distribution, and reproduction in any medium, provided the original author and source are credited
Protocol status: Working
We use this protocol and it's working
Created: May 13, 2023
Last Modified: May 13, 2023
Protocol Integer ID: 81825
Abstract
cell pellet dounce homogenisation
Protocol materials
Reagentnano-WCatalog ##2018-5ML
Step 11
HEADER
HEADER
SAN:
SPEC No:
OPERATOR:

Dounce Homogenisation
Dounce Homogenisation
  1. Remove cells from the flasks by scraping with a disposable plastic scraper and poor into a Amount10 mL centrifuge tube.

Pellet the cells in a bench top centrifuge at Centrifigation2000 rpm Duration00:02:00 , remove supernatant.

2m
Resuspend the cells in equal volume TC water and transfer to glass homogeniser tube.
Insert teflon plunger into cordless drill, set drill on max speed.
Homogenise(20 strokes) inside Class II BSC cabinet.
Clarify resultant solution at Centrifigation13000 rpm in Eppendorf centrifuge for Duration00:01:00


1m
Stand for Duration00:05:00 to permit viruses to diffuse back into solution, from the debris, and provide an interface for the sampling of membrane associated viruses. Use supernatant for sample below.

5m
Conventional
Conventional
11m
11m
Adsorb Amount10 µL sample to grid Duration00:10:00 , inspect to ensure sample does not dry out.

10m
Drain excess sample from grid using filter paper, leave wet.
Stain Reagentnano-WCatalog ##2018-5ML Duration00:01:00

1m
Drain & dry using filter paper