A procedure for microbial cell density determination with a high-throughput densitometric assay was developed to allow a precise quantification of both free and sessile cells, such as those of a biofilm, with a large range from low to high cell densities. Densitometry was chosen because it allows fast, rapid and cost-effective measures; it is non-disruptive; and has an easy learning curve. The method setup, and the further validation, was carried out with strains of Candida albicans, C. tropicalis and C. parapsilosis. Equations were developed at the level of the single strains, of the three species and finally a general one applicable to all three species. In the cross validation, with strains absent from the training set, the method was shown to be robust and flexible. The best results were obtained with species specific equations, although the global equation performed almost as well in terms of correlation between real and estimated density values. In all cases, a correlation around 0.98 between effective and predicted density was obtained with figures ranging from 102 to 108 cells mL−1. The entire analytical part of the procedure can be accomplished with a MS Excel macro provided free of charge.
High-Throughput Rapid and Inexpensive Assay for Quantitative Determination of Low Cell-Density Yeast Cultures
Debora Casagrande Pierantoni;Laura Corte;Luca Roscini;Gianluigi Cardinali
2019
Abstract
A procedure for microbial cell density determination with a high-throughput densitometric assay was developed to allow a precise quantification of both free and sessile cells, such as those of a biofilm, with a large range from low to high cell densities. Densitometry was chosen because it allows fast, rapid and cost-effective measures; it is non-disruptive; and has an easy learning curve. The method setup, and the further validation, was carried out with strains of Candida albicans, C. tropicalis and C. parapsilosis. Equations were developed at the level of the single strains, of the three species and finally a general one applicable to all three species. In the cross validation, with strains absent from the training set, the method was shown to be robust and flexible. The best results were obtained with species specific equations, although the global equation performed almost as well in terms of correlation between real and estimated density values. In all cases, a correlation around 0.98 between effective and predicted density was obtained with figures ranging from 102 to 108 cells mL−1. The entire analytical part of the procedure can be accomplished with a MS Excel macro provided free of charge.I documenti in IRIS sono protetti da copyright e tutti i diritti sono riservati, salvo diversa indicazione.
