Optimizing the recovery of peripheral blood mononuclear cells trapped in leukoreduction filters - A comparison study

Abstract Introduction The isolation of captured peripheral blood mononuclear cells (PBMNCs) from leukoreduction filters (LRFs) can be of great importance in terms of bringing the lost cells back into use. Objective The aim of this study was to evaluate various methods based on their potential to recover the peripheral blood cells from LRFs with a focus on mononuclear cells (MNCs). Method For cell isolation from LRFs, three distinct methods (back-flushing, direct and vacuum pump) were compared through the calculation of the yield of isolated MNCs. The viability of extracted cells was determined by the flow cytometry technique. Moreover, the recovered MNCs were characterized regarding the presence of blood stem cell purification. The cell culture, microscopic observation, and immunophenotyping were employed to characterize the blood stem cells (hematopoietic, mesenchymal and progenitor endothelial stem cells). Results The yield of isolation obtained in the back-flushing, direct and vacuum pump methods were 17.7 ± 1.28, 17.3 ± 0.96 and 21.2 ± 0.90 percent, respectively. Although the highest potential for total blood cell recovery belonged to the vacuum pump method, the lowest cell viability (85.73 ± 4.84%) was observed in this method. However, the isolation process of the back-flushing and direct methods had less effect on cell viability. The characterization of the isolated MNCs displayed that the dominant positive phenotype was for CD34/CD45, indicating hematopoietic stem cells. In addition, the endothelial stem/progenitor cells were significantly detected as CD31/CD133 positive cells. Conclusion According to our results and considering the safety and efficiency potential of each of the applied methods, the back-flushing in comparison with the other methods can be considered a suitable procedure for MNC isolation from LRFs.

Evaluation of Isolation and Identification Methods of Exosomes from Different Biofluids / 中国生物化学与分子生物学报

Exosomes are phospholipid bilayer extracellular vesicles secreted by cells, and they act as carriers for material transfer and information exchange between cells. Exosomes exist in many different biofluids and have good application prospects in disease diagnosis and drug delivery. Due to the nanoscale size and heterogeneity of exosomes, as well as the complex composition of biofluids, the separation of exosomes from biofluids is particularly difficult. At present, there are six commonly used methods for isolating exosomes from biofluids: ultracentrifugation, precipitation, size exclusion chromatography, density gradient centrifugation, ultrafiltration and immuno-affinity membrane. Each method has its own advantages and disadvantages. It is difficult to separate and obtain high-quality exosomes when used alone. There are also differences in the selection of methods for separating exosomes from different biofluids. With the continuous emergence of some new technologies such as microfluidic technology and the continuous improvement of classical methods, the exosome separation technology is improved, which is beneficial to the application of exosomes in clinical and scientific research. This review compares and evaluates the isolation and identification methods of exosomes from different biofluid sources, summarizes the selection and differences of exosome isolation methods from different biofluid sources, and provides a reference for subsequent research on exosomes from biofluids.

Research strategies for endophytes in medicinal plants based on high-throughput sequencing and traditional culture and isolation methods / 中国中药杂志

The research on endophytes of medicinal plants mainly relies on the traditional culture and isolation methods. Because of their functions such as promoting host growth, improving stress resistance, promoting the accumulation of medicinal active ingredients or directly producing medicinal active ingredients, the endophytes of medicinal plants have gradually attracted wide attention. However, it was found that the strains isolated by traditional methods were not the true dominant endophytes of medicinal plants by comparing the results of traditional culture isolation with high-throughput sequencing. The blind and random nature of traditional methods leads to the lack of standards in terms of medium selection, culture time and interaction between species. On the contrary, high-throughput sequencing technology is an emerging molecular biology technology developed in recent decades. Due to its high resolution level and indepen-dent culture, it can be used for thorough analysis of the community structure and diversity of environmental microorganisms. Therefore, we proposed the strategy of using high-throughput sequencing technology to guide the traditional culture and isolation of endophytes from medicinal plants. Firstly, the endophytic structure and diversity of medicinal plants were completely clear by high-throughput sequencing technology, and the dominant endophytes of the host were unequivocal. Then according to the characteristics of each dominant endophytes design or query suitable medium for its growth to culture and isolation. Finally, the function of the isolates was studied. This method can prevent researchers from missing out on the important functional strains of the host, expand the research scope of endophytes of medicinal plants, and facilitate the in-depth excavation and utilization of endophytes of medicinal plants.

Isolation of Candida albicans among HIV Patients Presenting with Oral Thrush in Abuja, Nigeria

Background: Candida infections are known contributors to the high morbidity and mortality rates seen in HIV positive patients.Methodology: This was a descriptive cross-sectional study. The study was carried out at the Microbiology research laboratory, department of Medical Microbiology, University of Abuja Teaching Hospital, Gwagwalada. The population is made up of two hundred and ten (210) patients who presented with oral thrush between fifteen years and seventy years which comprises of 160 HIV seropositive and 50 non age and sex matched HIV seronegative patients. Culture, Microscopy and ELISA methods were used for isolation of Candida albicans. An interviewer-administered, structured questionnaire was used as the study tool Results: The mean age for the isolation of Candida albicanswas 30 ± 18.7 years, with the highest proportion of isolates within the age range of 21-30 years accounting for 27.1% of the study population recruited and the lowest proportion of isolates being 41-50 years accounting for 8.0%.From the 210 subjects withCandida albicansin the study, one hundred and sixty (160) representing 76.2 were isolated from HIV seropositive clients and Candida albicansisolation rate among HIV seronegative population was 23.8%.Conclusion: The sensitivity, specificity and positivepredictive value of using Grams reaction methods in the diagnosis of Candida albicanswas 22.9%, 95.2% and 82.6%. The sensitivity, specificity and positive predictive value of using ELISA methods was 25.7%, 86.7% and 65.9%.In this study, there was preponderance of Candida albicansisolate among the young and the old in HIV seropositive patients but largely isolated from older patients among HIV seronegative patients

Some Approaches for the Selective Isolation of Rare Actinomycetes / 微生物学通报

The focus of microbiologists has moved to the rare actinomycetes.For selective isolation of rare actinomycetes that all play the important role in bioactive compounds,the approaches which involve the methods using gellan gum and flooding solution、 rehydration-centrifugation(RC)、 extremely high frequency radiation(EHF)、 bacteriophage and sucrose-gradient centrifugation were introduced in this paper.