[The existence of contamination in enzymes used for the isolation of Aspergillus and Candida species DNA]. / Aspergillus ve Candida cinsi DNA'sinin izolasyonunda kullanilan enzimlerde kontaminasyon varliginin arastirilmasi.

Molecular biological methods which are widely used in different fields, have been replaced with conventional diagnostic tests for the early diagnosis of invasive fungal infections, recently. Polymerase chain reaction (PCR) is one of these methods, with high specificity and sensitivity, which is accepted throughout the world. However, the enzymes that are used for the isolation of target DNA, may be contaminated with the gene sequences of some other fungal species in the preparation steps and may affect the PCR results. The studies showed that, the contamination of these enzymes with any type of fungi during their production steps, leads false positive results in PCR tests. So, the additional studies are recommended for minimizing the contamination. The aim of this study was to search whether the enzymes necessary for the isolation of fungal target DNA, used in PCR, are contaminated or not. For this purpose, five different enzymes namely, Zymolase 20T (from Arthrobacter luteus, two examples from different companies), lyticase (from Arthrobacter luteus), lysing enzyme (from Trichoderma harzianum) and proteinase K (from Tritrachium album) have been investigated for the presence of contamination. As a result, both of zymolase 20T enzymes were found to be contaminated from some fungal species with the demonstration of 18S rRNA gene sequences, while the other enzymes were found non-contaminated. By the help of this method, the most suitable enzyme for PCR was chosen and fungal contamination was prevented.

Successful clinical islet isolation using a GMP-manufactured collagenase and neutral protease.

In 2007, the islet community was notified that the collagenase product most commonly used for human islet isolations contained bovine neural tissue contaminants. To minimize this potential hazard, we adapted our human islet processing procedure to use a GMP-manufactured, bovine neural tissue-free collagenase blend. Here, we describe the factors that we consider most important for achieving reproducible and clinically useable islet isolations using this product.

Separation of proteolytic enzymes originating from Antarctic krill (Euphausia superba) by capillary electrophoresis.

Extracts prepared from Antarctic krill (Euphausia superba), mainly consisting of acidic proteolytic enzymes, have been studied with capillary electrophoretic techniques. Approximately 50 repeatable peaks were obtained with capillary zone electrophoresis on an untreated fused-silica capillary using a phosphate buffer containing anionic and cationic fluorosurfactant additives as separation medium. A faster separation was achieved on a polyvinyl alcohol coated capillary. Quantitative variations of individual proteins regarding different krill enzyme batches were noted. In the krill samples trypsin-like serine proteinase, carboxypeptidase A and carboxypeptidase B were tentatively identified.