Cell membrane rupture: a novel test reveals significant variations among different brands of tissue culture flasks

Objectives Loss of cytoplasmic molecules including protein controls, due to cell membrane rupture can cause errors and irreproducibility in research data. Previous results have shown that during the washing of a monolayer of cells with a balanced salt solution, the fluid force causes cell membrane rupture on some areas of the flasks/dishes. This fact shows the non-uniformity of the polystyrene surface in terms of cell culture. There is at present no simple test to monitor that surface. This paper presents a novel biologically based assay to determine the degree of heterogeneity of flasks supplied by various manufacturers. Results This paper shows that significant variation exists in polystyrene surface heterogeneity among several brands of tissue culture flasks, varying from 4 to 20% of the flask surface. There is also large variability within the production lot of a manufacturer. The assay method involves loading the cells with a cytoplasmic fluorescent marker that is released upon cell membrane rupture. Cell membrane rupture also causes the loss of marker proteins such as GAPDH used in Westernblots. This novel assay method can be used to monitor the batch consistency and the manufacturing process of flasks/dishes. It may also be used to test new biomaterials.

Solvent- and Initiator-free Fabrication of Efficient and Stable Perovskite-Polystyrene Surface-Patterned Thin Films for LED Backlights

We report a one-pot route for synthesis of CsPbBr3 perovskite nanocrystals (PNCs) in styrene to form glue-like polystyrene(PS) pre-polymer incorporating mono-dispersed PNCs. The pre-polymer enables solvent- and initiator-free fabricating and...

Lactic Acid Bacteria against Listeria monocytogenes

Background: Listeria monocytogenes is a pathogenic bacterium that can contaminate food and cause public health problems due its ability to form biofilms and resistance to sanitizers, it is responsible for sanitary and economic losses in food producing establishments. The difficulties in controlling biofilms and increasing resistance to traditional antibacterial agents is motivating studies of alternative potential biological agents for the control of pathogenic biofilms, among which lactic acid bacteria (LABs) are included. The objective of this work was to evaluate the activity of LABs against Listeria monocytogenes biofilm formation on polystyrene plates, a surface commonly used in the food industry.Materials, Methods & Results: Lyophilized commercial strains of Bifidobacterium animalis, Lactobacillus fermentum, Lactobacillus plantarum, Lactobacillus salivaris and Lactobacillus acidophilus were used. The strain of Listeria monocytogenes (L4) was isolated from polystyrene mats from a poultry slaughterhouse cutting room and demonstrated the ability to attach to microplates and resistance to sanitizers (sodium hypochlorite and hydrogen peroxide) at all times, temperatures and tested surfaces. The antimicrobial activity of LABs was evaluated by the agar diffusion method. The LABs that presented action on Listeria monocytogenes were selected for the inhibition and/or removal of biofilms in microplates, and all experiments were carried out in triplicate. Only Bifidobacterium animalis and Lactobacillus plantarum demonstrated action against Listeria. monocytogenes in the agar diffusion assays and were selected for inhibition and competition assays. Furthermore, competition of LABs against Listeria monocytogenes adhesion was evaluated. There was no significant difference between LABs and Listeria monocytogenes, alone or in combination, at temperatures of 30ºC and 37ºC in the Listeria monocytogenes inhibition assays on polystyrene surface. The lactic acid bacteria evaluated did not demonstrate inhibition of Listeria monocytogenes adhesin testes with optical density visualization, however, it was possible to identify a reduction in Listeria monocytogenes counts with the application of Bifidobacterium animals and Lactobacillus plantarum in the testes of competition against biofilm formation. In competition tests Bifidobacterium animalis and Lactobacillus plantarum have an injunction in Listeria monocytogenes, indicating that these lactic acid bacteria can retard Listeria biofilm formation on polystyrene surfaces and thus help control the pathogen in the food industry.Discussion: A potential mechanism to control biofilm adhesion and formation of pathogens for nutrients and fixation on surfaces, multiplication factors and surfaces are a challenge in controlling biofilms of pathogenic microorganisms, alternative measures to traditional methods for inactivating pathogens and biofilm formers bacteria are necessary. In this sense, lactic acid bacteria generate high levels of bacteriocin and are effective in inhibiting the biofilm of pathogenic bacteria, however, our study did not reveal this. We verified that Bifidobacterium animalis and Lactobacillus plantarum have an inhibitory action on Listeria monocytogenes, indicating that these lactic acid bacteria can be used to delay the formation of biofilms by Listeria on polystyrene surfaces, helping to control this pathogen in food industry.Keywords: control of biofilm, pathogenic bacteria, food industry, polystyrene surface, FTDs.

Cell membrane rupture: A novel test reveals significant variations among different brands of tissue culture flasks

Objectives Loss of cytoplasmic molecules including protein controls, due to cell membrane rupture can cause errors and irreproducibility in research data. Previous results have shown that during the washing of a monolayer of cells with a balanced salt solution, the fluid force causes cell membrane rupture on some areas of the flasks/dishes. This fact shows the non-uniformity of the polystyrene surface in terms of cell culture. There is at present no simple test to monitor that surface. This paper presents a novel biologically based assay to determine the degree of heterogeneity of flasks supplied by various manufacturers. Results This paper shows that significant variation exists in polystyrene surface heterogeneity among several brands of tissue culture flasks, varying from 4% to 20% of the flask surface. There is also large variability within the production lot of a manufacturer. The assay method involves loading the cells with a cytoplasmic fluorescent marker that is released upon cell membrane rupture. Cell membrane rupture also causes the loss of marker proteins such as GAPDH used in Westernblots. This novel assay method can be used to monitor the batch consistency and the manufacturing process of flasks/dishes. It may also be used to test new biomaterials.

Cell Membrane Rupture: A Novel Test Reveals Significant Variations Among Different Brands of Tissue Culture Flasks

Unanticipated errors in scientific research data can be attributed to the unwarranted assumption of uniformity in the polystyrene surface that is ubiquitously used in tissue culture flasks and dishes. We have shown that when adherent cells are subjected to fluid shear force, equivalent to rinsing the culture with a balanced salt solution, cells on some areas of the polystyrene surface will immediately rupture while still adherent on the surface. This heterogeneity on the polystyrene surface can cause unexpected variability in experimental results and in replicating experiments among labs. In this paper a novel quantitative method is described to measure the degree of heterogeneity on the polystyrene surface of tissue culture flasks. The results show significant variation among several brands of tissue culture flasks as well as large variability within the production lot of a manufacturer. The assay method involves loading the cells with a fluorescent marker that is released upon membrane rupture. Cell membrane rupture also causes the loss of marker proteins used in Westernblots. This novel assay method can be used to monitor the batch consistency and the manufacturing process of flasks and dishes. It may also be used to test new biomaterials.