scholarly journals An Improved Crystal Violet Assay for Biofilm Quantification in 96-Well Microtitre Plate

2017 ◽  
Author(s):  
Sudhir K. Shukla ◽  
T. Subba Rao
Keyword(s):  
Edge Effect ◽  
High Throughput ◽  
Crystal Violet ◽  
High Throughput Screening ◽  
Experimental Error ◽  
Rejection Rate ◽  
Microtitre Plate ◽  
Crystal Violet Assay ◽  
Biofilm Quantification ◽  
Classical Crystal

AbstractMicroplates are essential tools for biofilm research since it allows high throughput screening of biofilm forming strains or in the assay of anti-biofilm drugs. However, 96 well microtitre plate based assays share the issue of “edge effect”. The primary cause of the “edge effect” phenomenon is evaporation. As edge effect causes a significant increase in plate rejection rate by introducing experimental error, we improvised the classical crystal violet assay to reduce water loss from the peripheral wells. The improvised method showed a significant reduction in edge effect and minimised error in crystal violet assay

scholarly journals A Simple Technique for Reducing Edge Effect in Cell-Based Assays

2003 ◽  
Vol 8 (5) ◽  
pp. 566-570 ◽  
Author(s):  
Betina Kerstin Lundholt ◽  
Kurt M. Scudder ◽  
Len Pagliaro
Keyword(s):  
Edge Effect ◽  
High Throughput ◽  
Edge Effects ◽  
High Throughput Screening ◽  
Room Temperature ◽  
Simple Technique ◽  
Rejection Rate ◽  
Uneven Distribution ◽  
Ambient Conditions ◽  
Number Of Factors

Several factors are known to increase the noise and variability of cell-based assays used for high-throughput screening. In particular, edge effects can result in an unacceptably high plate rejection rate in screening runs. In an effort to minimize these variations, the authors analyzed a number of factors that could contribute to edge effects in cell-based assays. They found that pre-incubation of newly seeded plates in ambient conditions (air at room temperature) resulted in even distribution of the cells in each well. In contrast, when newly seeded plates were placed directly in the CO2 incubator, an uneven distribution of cells occurred in wells around the plate periphery, resulting in increased edge effect. Here, the authors show that the simple, inexpensive approach of incubating newly seeded plates at room temperature before placing them in a 37° C CO2 incubator yields a significant reduction in edge effect. ( Journal of Biomolecular Screening 2003:566-570)

scholarly journals Adaptation of the Start-Growth-Time Method for High-Throughput Biofilm Quantification

2021 ◽  
Vol 12 ◽  
Author(s):  
Lara Thieme ◽  
Anita Hartung ◽  
Kristina Tramm ◽  
Julia Graf ◽  
Riccardo Spott ◽  
...  
Keyword(s):  
High Throughput ◽  
High Throughput Screening ◽  
Growth Curves ◽  
Antimicrobial Effect ◽  
Growth Time ◽  
Colony Forming Unit ◽  
Resazurin Assay ◽  
Unspecific Binding ◽  
Biofilm Quantification ◽  
Doubling Times

Colony forming unit (CFU) determination by agar plating is still regarded as the gold standard for biofilm quantification despite being time- and resource-consuming. Here, we propose an adaption of the high-throughput Start-Growth-Time (SGT) method from planktonic to biofilm analysis, which indirectly quantifies CFU/mL numbers by evaluating regrowth curves of detached biofilms. For validation, the effect of dalbavancin, rifampicin and gentamicin against mature biofilms of Staphylococcus aureus and Enterococcus faecium was measured by accessing different features of the viability status of the cell, i.e., the cultivability (conventional agar plating), growth behavior (SGT) and metabolic activity (resazurin assay). SGT correlated well with the resazurin assay for all tested antibiotics, but only for gentamicin and rifampicin with conventional agar plating. Dalbavancin treatment-derived growth curves showed a compared to untreated controls significantly slower increase with reduced cell doubling times and reduced metabolic rate, but no change in CFU numbers was observed by conventional agar plating. Here, unspecific binding of dalbavancin to the biofilm interfered with the SGT methodology since the renewed release of dalbavancin during detachment of the biofilms led to an unintended antimicrobial effect. The application of the SGT method for anti-biofilm testing is therefore not suited for antibiotics which stick to the biofilm and/or to the bacterial cell wall. Importantly, the same applies for the well-established resazurin method for anti-biofilm testing. However, for antibiotics which do not bind to the biofilm as seen for gentamicin and rifampicin, the SGT method presents a much less labor-intensive method suited for high-throughput screening of anti-biofilm compounds.

scholarly journals Inhibitory effects of antifungal drugs on biofilm producing Aspergillus spp. recovered from drinking water system

2020 ◽  
Author(s):  
Noorulain Nazir ◽  
Abubakar Siddique ◽  
Nisar A Khan
Keyword(s):  
Drinking Water ◽  
Amphotericin B ◽  
Crystal Violet ◽  
Biofilm Formation ◽  
Respiratory Diseases ◽  
Antifungal Susceptibility ◽  
Antifungal Drugs ◽  
Microtitre Plate ◽  
Plate Method ◽  
Crystal Violet Assay

Abstract Aims: Biofilms formed in drinking water distribution systems serve as a continuous source of fungal infections. Biofilms are thick aggregates of adherent microorganisms including pathogenic species of fungi. Respiratory diseases and skin allergy reactions are caused by drinking water containing biofilm forming fungus and bacteria. One of the main causes of nosocomial infections and respiratory diseases in hospitals is due to the fungal biofilm formation in machines, catheters and other surgical instruments. There are some antifungal drugs which are used to control biofilm formation to minimize the infection rate. Methodology and results: The present study was conducted to isolate and identify Aspergillus species which are the main fungal spp. responsible for the biofilm formation in drinking water and to check their antifungal susceptibility against antifungal drugs. The isolated fungal samples from drinking water were cultivated on Potato dextrose agar for the isolation of Aspergillus species. Isolated Aspergillus species were identified on the basis of cultural, morphological and microscopic examination. Then in-vitro ability of biofilm produced by isolated Aspergillus species was estimated using microtitre plate method and quantification by crystal violet assay. Antifungal susceptibility testing against isolated fungal spp. was done by antifungal drug Amphotericin B.Results: From results, it is concluded that drinking water of labs, hospitals and common water chillers were more prevelant by Aspergillus species whereas water from reverse osmosis plants showed negative results. From microtitre plate method and crystal violet assay, it was concluded that Aspergillus spp. are Susceptible against Amphotericin B drug as compared to miconazole.

scholarly journals High-throughput screening alternative to crystal violet biofilm assay combining fluorescence quantification and imaging

2021 ◽  
pp. 106343
Author(s):  
Cristina Isabel Amador ◽  
Rune Overlund Stannius ◽  
Henriette Lyng Røder ◽  
Mette Burmølle
Keyword(s):  
High Throughput ◽  
Crystal Violet ◽  
High Throughput Screening

High-throughput screening program for botanical extracts

Planta Medica ◽  
2012 ◽  
Vol 78 (11) ◽  
Author(s):  
L Hingorani ◽  
NP Seeram ◽  
B Ebersole
Keyword(s):  
High Throughput ◽  
High Throughput Screening ◽  
Screening Program ◽  
Botanical Extracts

High throughput screening of microbial biodiversity for the discovery of novel cosmeceutical agents

Planta Medica ◽  
2015 ◽  
Vol 81 (16) ◽  
Author(s):  
K Georgousaki ◽  
N DePedro ◽  
AM Chinchilla ◽  
N Aliagiannis ◽  
F Vicente ◽  
...  
Keyword(s):  
High Throughput ◽  
High Throughput Screening ◽  
Microbial Biodiversity

High throughput screening to investigate Brazilian Cerrado biome plant extract bank chemical diversity

Planta Medica ◽  
2016 ◽  
Vol 81 (S 01) ◽  
pp. S1-S381
Author(s):  
LS Espindola ◽  
RG Dusi ◽  
KR Gustafson ◽  
J McMahon ◽  
JA Beutler
Keyword(s):  
High Throughput ◽  
Plant Extract ◽  
High Throughput Screening ◽  
Chemical Diversity ◽  
Brazilian Cerrado ◽  
Cerrado Biome

High-throughput screening revealed a clinically relevant drug to induce sperm motility

2014 ◽  
Author(s):  
Clair Cochrane ◽  
Halil Ruso ◽  
Anthony Hope ◽  
Rosemary G Clarke ◽  
Christopher Barratt ◽  
...  
Keyword(s):  
Sperm Motility ◽  
High Throughput ◽  
High Throughput Screening
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