Precise and High-Throughput Analysis of Bacterial Growth Using a Microplate Reader

Abstract

Source: Kurokawa, M., et al. Precise, High-throughput Analysis of Bacterial Growth. J. Vis. Exp. (2017)

This video demonstrates the high-throughput quantification of bacterial growth using a microplate reader. It outlines the steps involved in culture preparation, optical density measurement, and analysis of growth dynamics.

Protocol

1. Real-time recording of growth
   1. Draw a 96-well plate pictogram (8 × 12 table) to indicate the positions of the inoculated culture samples on the 96-well plate. Print out the table and use it as a reference for the experiment.
      NOTE: The wells located on the edges of the microplate should only contain blank medium because of evaporation.
   2. Place a sterilized 96-well flat-bottom microplate with lid, P-1000, 1000-µL pipette tips, P-200, 200-µL pipette tips, several 1.5-mL microtubes, an 8-multichannel pipette, a sterilized reagent reservoir, room temperature M63, and the Escherichia coli<....

Materials

List of materials used in this article

Name	Company	Catalog Number	Comments
Dipotassium phosphate	Wako	164-04295
Monopotassium phosphate	Wako	166-04255
Ammonium sulfate	Wako	019-03435
Magnesium sulfate heptahydrate	Wako	138-00415
Thiamine-HCl	Wako	201-00852
Glucose	Wako	049-31165
HCl	Wako	080-01066
Iron (II) sulfate heptahydrate	Wako	094-01082
KOH	Wako	168-21815
Glycerol	Wako	075-00611
Pipette Tips, 200 µL	WATSON	110-705Y
Pipette Tips, 1,000 µL	WATSON	110-8040
Microtube (1.5 mL)	WATSON	131-715C
8 multichannel-pipette	WATSON	NT-8200
Pasorina stirrer	AS ONE	2-4990-02
Glass cylinder (200 mL)	AS ONE	1-8562-07
Precision pH meter	AS ONE	AS800 / 1-054-01
Pipetman P-200	GILSON	1-6855-05
Pipetman P-1000	GILSON	1-6855-06
Microtube stand	BM Bio	801-02Y
Vortex	BM Bio	BM-V1
Corning Costar 96-well microplate with lid (Flat bottom, Clear)	Sigma-Aldrich	Corning, 3370
Corning Costar reagent reservoir (50 mL)	Sigma-Aldrich	Corning, 4870
EPOCH2	BioTek	2014-EP2-002 / EPOCH2T
BIO clean bench	Panasonic	MCV-B131F
Bacterial strains	Strain bank organization; National Bio Resource Project (NBRP) in Japan