Simple detection of small amounts of Pseudomonas cells in milk by using a microfluidic device.
OBJECTIVE
Flow cytometry offers rapid and reliable analyses of bacteria in milk. However, a flow cytometer is relatively expensive and operation is rather complicated for an unskilled operator. We applied flow cytometry using a microfluidic device (on-chip flow cytometry) in detection of small amounts of milk-spoiling bacteria.


RESULTS
Pseudomonas cells in milk were in situ hybridized with Cy5-labelled probe specific for Pseudomonas spp. under optimized condition. Numbers of Pseudomonas cells in the stationary phase and in the starved state determined by on-chip flow cytometry were compared with those determined by conventional plate counting, and on-chip flow cytometry detected targeted cells in milk that were undetectable as colony forming units(CFU) on Standards Methods Agar.


CONCLUSIONS
The contamination in milk with fewer than 10 CFU ml(-1) of targeted cells in starved state was detectable with simple procedure (0.5 h milk-clearing, 1 h fixation, 2 h hybridization and 0.5 h on-chip flow cytometry following 12 h enrichment of cells).


CONCLUSIONS
On-chip flow cytometry following fluorescence in situ hybridization could be applicable to simple detection of milk-spoiling bacteria.
Yamaguchi
N
N
Graduate School of Pharmaceutical Sciences, Osaka University, Suita, Osaka, Japan.
Ohba
H
H
Nasu
M
M
D000818:Animals; D015169:Colony Count, Microbial; D005434:Flow Cytometry / Q000379:methods*; D005516:Food Microbiology; D017404:In Situ Hybridization, Fluorescence / Q000379:methods*; D046210:Microfluidic Analytical Techniques / Q000379:methods*; D008892:Milk / Q000382:microbiology*; D011549:Pseudomonas / Q000235:genetics / Q000254:growth & development / Q000302:isolation & purification*