ABSTRACT
The mitochondria toxicity assay (MTT assay) is an established method for monitoring cell viability based on mitochondrial activity. Here the MTT assay is proposed for the in situ quantification of the living cell density of microencapsulated Jurkat cells. Three systems were used to encapsulate the cells, namely a membrane consisting of an interpenetrating polyelectrolyte network of sodium cellulose sulphate/poly(diallyldimethylammonium chloride) (NaCS/PDADMAC), a calcium alginate hydrogel covered with poly(L-lysine) (Ca-alg-PLL), and a novel calcium alginate-poly(ethylene glycol) hybrid material (Ca-alg-PEG). MTT results were correlated to data obtained by the trypan blue exclusion assay after release of the cells from the NaCS/PDADMAC and Ca-alg-PLL capsules, while a resazurin-based assay was used for comparison in case of the Ca-alg-PEG material. Analysis by MTT assay allows quick and reliable determination of viable cell densities of encapsulated cells independent of the capsule material. The assay is highly reproducible with inter-assay relative standard deviations below 10%.
Subject(s)
Alginates/pharmacology , Cellulose/analogs & derivatives , Mitochondria/drug effects , Polyethylene Glycols/pharmacology , Polyethylenes/pharmacology , Quaternary Ammonium Compounds/pharmacology , Cell Survival/drug effects , Cellulose/pharmacology , Glucuronic Acid/pharmacology , Hexuronic Acids/pharmacology , Humans , Jurkat Cells , Structure-Activity RelationshipABSTRACT
An amperometric enzyme-based sensor-system for the direct detection of formaldehyde in air is under investigation. The biosensor is based on a native bacterial NAD(+)- and glutathione-independent formaldehyde dehydrogenase as biorecognition element. The enzyme was isolated from Hyphomicrobium zavarzinii strain ZV 580, grown on methylamine hydrochloride in a fed-batch process. The sensor depends on the enzymatic conversion of the analyte to formic acid. Released electrons are detected in an amperometric measurement at 0.2V vs. Ag/AgCl reference electrode by means of a redox-mediator. To optimize the sensing device, Ca(2+) and pyrroloquinoline quinone (PQQ) were added to the buffer solution as reconstitutional substances. At this stage, the sensor shows linear response in the tested ppm-range with a sensitivity of 0.39 microA/ppm. The signal is highly reproducible with respect to sensitivity and base line signal. Reproducibility of sensitivity is more than 90% within the same bacterial batch and even when enzyme of different bacterial batches is used.
