The Direct Epifluorescent Filter Technique (DEFT)

2003 ◽  
pp. 113-122
Author(s):  
Rohan Kroll
Keyword(s):  
Filter Technique ◽  
Direct Epifluorescent Filter Technique

Determination of Viable Bacterial Populations in Raw Milk within 20 Minutes by Using a Direct Epifluorescent Filter Technique

1997 ◽  
Vol 60 (7) ◽  
pp. 874-876 ◽  
Author(s):  
CLAUDE P. CHAMPAGNE ◽  
NANCY J. GARDNER ◽  
JULIE FONTAINE ◽  
JACQUES RICHARD
Keyword(s):  
Raw Milk ◽  
Plate Count ◽  
Bacterial Populations ◽  
Filter Technique ◽  
Milk Samples ◽  
Standard Plate ◽  
Plate Counts ◽  
Direct Epifluorescent Filter Technique ◽  
Triton X

The results from a shortened procedure for the direct epifluorescent filter technique (DEFT) determination of viable bacterial populations in raw milk were compared to standard plate counts. Shortening the prefiltration trypsin-Triton X-100 incubation period from 10 to 3 min enabled the completion of the analysis within 20 min. The short DEFT method results had a correlation coefficient (r) of 0.81 with plate counts. With respect to precision, the average difference between values of duplicate plate count analyses was 0.16 log units; that of the short DEFT was 0.14 log units. The slopes of the regressions equations were less than 1, indicating that a direct correlation is not achieved. Short DEFT values were 0.17 log units higher than those of plate counts on milk samples containing less than 10,000 CFU/ml. For milk samples containing counts over 10,000 CFU/ml, short DEFT values averaged only 0.05 log units above plate count readings. Daily preparation of the stain appears unnecessary since acridine orange solutions stored for up to 2 days at 4°C did not produce results significantly (P > 0.05) different from those obtained with fresh solutions. The short DEFT method has potential for the assessment of the bacteriological quality of raw milk in tanker deliveries.

The Limulus Amoebocyte Lysate Assay and the Direct Epifluorescent Filter Technique as Estimators of Potential Shelf-life of Pasteurized Fluid Milk

1990 ◽  
Vol 53 (2) ◽  
pp. 151-153 ◽  
Author(s):  
ROBERT D. BYRNE ◽  
J. RUSSELL BISHOP
Keyword(s):  
Shelf Life ◽  
Benzalkonium Chloride ◽  
Detection Techniques ◽  
Filter Technique ◽  
Psychrotrophic Bacteria ◽  
Limulus Amoebocyte Lysate ◽  
Whole Milk ◽  
Before And After ◽  
Direct Epifluorescent Filter Technique ◽  
Fluid Milk

The Limulus Amoebocyte Lysate Assay, Direct Epifluorescent Filter Technique and modified Psychrotrophic Bacteria Count were used to indicate potential shelf-life of pasteurized fluid milk. Commercial whole milk samples, stored at 7°C, were analyzed for bacterial and biochemical parameters, as well as for potential shelf-life by daily sensory evaluation. Each sample was evaluated before and after the following preliminary incubations: milk alone, milk with benzalkonium chloride, milk and broth, and milk and broth with benzalkonium chloride. The Limulus Amoebocyte Lysate Assay, Direct Epifluorescent Filter Technique, and modified Psychrotrophic Bacteria Count in conjunction with the preliminary incubations, produced relatively high correlations to shelf-life (−0.78, −0.85, and −0.86 respectively). Thus, these bacterial detection techniques could be used as rapid methods of shelf-life estimation.

Effect of Agitation on Bacterial Aggregates in Pure Cultures and Raw Milk

1983 ◽  
Vol 46 (8) ◽  
pp. 681-685 ◽  
Author(s):  
ROBYN E. O'CONNOR ◽  
K. N. EWINGS ◽  
NEIL W. HOLLYWOOD
Keyword(s):  
Raw Milk ◽  
Mechanical Agitation ◽  
Filter Technique ◽  
Milk Samples ◽  
Pure Cultures ◽  
Plate Counts ◽  
Bacterial Aggregates ◽  
Direct Epifluorescent Filter Technique ◽  
Clump Size

A comparison of the effects of various mechanical agitation treatments on bacterial aggregates was performed on 8 pure cultures and 27 raw milk samples. Although both syringing and blending produced significant increases in total counts and psychrotroph counts, blending for 2 min gave the greatest increase in count. Use of the direct epifluorescent filter technique (DEFT) confirmed that syringing and blending reduced bacterial clump size to approximately 2 cells. These agitation treatments markedly improved the correlation between DEFT counts and plate counts.

Evaluation of the direct epifluorescent filter technique for assessing the hygienic condition of milking equipment

1983 ◽  
Vol 50 (1) ◽  
pp. 9-16 ◽  
Author(s):  
Alastair C. Hunter ◽  
Rhona M. McCorquodale
Keyword(s):  
Heat Treatment ◽  
Correlation Coefficients ◽  
Testing Methods ◽  
Hygienic Condition ◽  
Filter Technique ◽  
Ringer’S Solution ◽  
Streptococcus Lactis ◽  
Bright Orange ◽  
Living Organisms ◽  
Direct Epifluorescent Filter Technique

SummaryThe hygienic condition of 6 milking installations, 3 sanitized by circulation cleaning (CC) with chlorine-based chemicals and 3 by flushing with acidified boiling water (ABW), was tested using rinses of quarter strength Ringer's solution. The bacterial content of the rinses was determined using both colony counts and the direct epifluorescent filter technique (DEFT). A comparison of testing methods gave correlation coefficients between colony count and DEFT of 0·82 for plants using CC and 0·46 for plants using ABW.Five strains of bacteria belonging to different genera and commonly found on milking equipment were exposed to various degrees of heat and to various concentrations of chlorine. The effects of such treatments on the staining characteristics of the organisms were studied. It was observed thatStaphylococcus aureusandStreptococcus lactis, although killed by heat treatment, stained a bright orange when treated with acridine orange dye.Pseudomonas fluorescens, Escherichia coliand vegetative cells ofBacillus cereusdid not take up the orange stain after heat treatment, nor did any of the 5 strains stain orange after treatment with NaOCl. It is suggested that the DEFT is a useful and rapid means of counting bacteria in rinses of equipment where sterilization is due primarily to chlorination, but in the absence of a stain which can differentiate more accurately between dead and living organisms its use is limited where sterilization is carried out solely by heat.

Antibody-Direct Epifluorescent Filter Technique and Immunomagnetic Separation for 10-h Screening and 24-h Confirmation of Escherichia coli O157:H7 in Beef

1996 ◽  
Vol 59 (10) ◽  
pp. 1072-1075 ◽  
Author(s):  
LAWRENCE RESTAINO ◽  
H. JEFFREY CASTILLO ◽  
DIANA STEWART ◽  
MARY LOU TORTORELLO
Keyword(s):  
Escherichia Coli ◽  
Membrane Filtration ◽  
Immunomagnetic Separation ◽  
Rapid Screening ◽  
Epifluorescence Microscopy ◽  
Objective Lens ◽  
Escherichia Coli O157 ◽  
E Coli ◽  
Filter Technique ◽  
Direct Epifluorescent Filter Technique

Rapid screening of beef for the presence of Escherichia coli O157:H7 was shown to be feasible using a 10-h enrichment in modified buffered peptone water and the antibody-direct epifluorescent filter technique (Ab-DEFT). The Ab-DEFT involved membrane filtration, fluorescent antibody staining and epifluorescence microscopy and was accomplished in less than 1 h. The procedure allowed detection of the pathogen artificially inoculated into beef patties at 0.1 CFU/g. The 10-h nonselective enrichment broth supported rapid growth, which provided sufficient numbers of cells for a positive determination by the Ab-DEFT after fewer than 10 microscope fields were scanned using a 40× objective lens. Immunomagnetic separation using anti-E. coli O157 Dynabeads® was used to confirm presumptively positive cultures within 24 h. The ease and rapidity of the Ab-DEFT may provide a substantial time and cost savings to the beef industry for screening beef for the presence of E. coli O157:H7.

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