Preparation of Immunomagnetic Beads Enrichment Kit for Detection of Aflatoxin B1

Immunomagnetic beads enrichment kit for detection of aflatoxin B1(AFB1) was prepared through reaction of AFB1 and p-phenylenediamine. The catches of AFB1 by the kit were 25 ng/mg. Furthermore, AFB1 was conducted specific reaction with competitive drugs with similar structure or function to AFB1, including aflatoxin M1, T-2 toxin, ochratoxin A, zearalenone and patulin, and no cross reaction was observed.

On-chip immunomagnetic bead swarm based on magnetic actuation and mechanical vibration for biological detection

Immunomagnetic bead(IMB)-based detection has great potential for biomedical applications.Passive and active strategies,including microfluidics and magnetic actuation methods,have been developed to mix IMBs and analytes efficiently.However,cost-effective on-site detection using a simple microfluidic chip is challenging,and miniaturization of the magnetic driving device is imperative for portability.In this study,we propose a novel mixing method for an on-chip IMB swarm via magnetic actuation and mechanical vibration.A microfluidic chip system coupled with double spiral magnetic coils and a vibration motor was fabricated.The aggregation behavior of IMBs under magnetic fields and the diffusion behavior of the IMB swarm under mechanical vibration were analyzed in detail.Based on the synergetic effects of magnetic actuation and mechanical vibration,we achieved the highly efficient capturing of Vibrio parahaemolyticus DNA and goat anti-human immunoglobulin G by mixing the IMB swarm with the microfluidic chip.In this case,the antigen detection rate could reach~94.4%.Given its fascinating features,such IMB-microfluidic detection demonstrates significant potential for biomedical applications.

Isolation,cultivation and identification of brain glioma stem cells by magnetic bead sorting

This study describes a detailed process for obtaining brain glioma stem cells from freshly dissected human brain glioma samples using an immunomagnetic bead technique combined with serum-free media pressure screening. Furthermore, the proliferation, differentiation and self-renewal biological features of brain glioma stem cells were identified. Results showed that a small number of CD133 positive tumor cells isolated from brain glioma samples survived as a cell suspension in serum-free media and proliferated. Subcultured CD133 positive cells maintained a potent self-renewal and proliferative ability, and expressed the stem cell-specific markers CD133 and nestin. After incubation with fetal bovine serum, the number of glial fibrillary acidic protein and microtubule associated protein 2 positive cells increased significantly, indicating that the cultured brain glioma stem cells can differentiate into astrocytes and neurons. Western blot analysis showed that tumor suppressor phosphatase and tensin homolog was highly expressed in tumor spheres compared with the differentiated tumor cells. These experimental findings indicate that the immunomagnetic beads technique is a useful method to obtain brain glioma stem cells from human brain tumors.

Study on Shigella Detection by ATP Bioluminescence Magnetic Enzyme Immunoassay

[Objective] The research aimed to establish a rapid detection method for Shigella. [Method] Combining immunomagnetic separation technology with ATP bioluminescence technology, a new kind of fast and accurate ATP bioluminescence magnetic enzyme immunoassay technique for Shigella was established. [Result] Using ATP bioluminescence magnetic enzyme immunoassay technique to detect standard solution for Shigella （ATCC 25931 ）, result showed that correlation coefficient between relative light intensity detected by instrument and bacteria concentration detec- ted by culture counting method was 0.981 1. Moreover, relation curve between relative light intensity and Shigella concentration was drawn. [ Conclusion] The method had a high detection speed and accuracy, and could be used for the rapid detection of pathogen in food and environment.

Separation of CD34 Positive Cells and Determination of Surface Homing Antigen

To study the differences in homing potential between bone marrow cells and umbilical blood cells, CD34 positive cells were obtained from bone marrow (BM) and umbilical blood (UB) by the direct cell separation with domestic immunomagnetic beads. The expression of the two adhesion molecules CD11a/CD18 and CD44 were examined. After separation, CD34 positive cells accounted for 51 %-82 % of the harvested cells and dye-resistance rate was 82 %-88 %. The expression of CD11a/CD18 and CD44 on the surfaces of UB cells was 49. 6 % 1 10. 2 % and 37. 7 % ± 10. 3 % respectively. On BM cells they were 50. 2 % ± 6. 2 % and 34 % ± 13. 3 % respectively. There were no significant differences in the expression of these two molecules. It was concluded .that the cell separation method with domestic immunomagnetic beads was effective and the stem cells from UB could serve as an alternative source for transplantation.

Development of Annexin V Technology for Removal of Dead Spermatozoa from Bovine Spermatozoa and Its Application in Frozen Sexed Semen Production

[Objectives]This study was conducted to develop a molecular marker immunomagnetic bead sorting technology method that can specifically identify dead spermatozoa.[Methods]This study first confirmed the specific binding of Annexin V to dead bovine spermatozoa,and tried to remove dead spermatozoa in semen combining with the immunomagnetic bead technology,so as to improve the separation efficiency of target spermatozoa in the process of sex-controlled semen preparation on a flow cytometer.[Results]The spermatozoon motility,membrane integrity and mitochondrial activity after sorting and the rate of dead spermatozoa during the on-machine X/Y separation were all improved to different degrees(P<0.05),indicating that the technical process design could effectively remove some dead spermatozoa,and there was no significant effect on frozen sexed semen prepared from the separated X or Y spermatozoa(P>0.05),indicating that the technical process did not cause additional damage to the spermatozoa.[Conclusions]Combining the specificity of Annexin V with the immunomagnetic bead method could effectively remove dead spermatozoa from bovine spermatozoa,and significantly reduce the rate of dead spermatozoa in bovine permatozoa during sex-controlled separation(P<0.05).The method developed can effectively improve the production efficiency of frozen sexed semen of dairy cows,reduce the production cost,and promote the industrial application of the product.

Nominal effective immunoreaction volume of magnetic beads at single bead level

Immunomagnetic bead（IMB）-based enzyme-linked immunosorbent assay（ELISA） has been the tool frequently used for protein detection in research and clinical laboratories.For most ELISA reactions the recommended dosage of IMBs is usually according to their weight（mg） or mass fraction（w/v） instead of the bead number.Consequently, the processes occurring in the immediate vicinity of the IMBs have always been ignored by researchers and they cannot be revealed in detail during the ELISA reaction.In this paper, we established the relationship between number of IMBs and colorimetric results, and further proposed a new concept of ＂nominal effective immunoreaction volume（NEIV）＂ to characterize a single IMB during ELISA reaction.Results showed that the NEIV of a single IMB has a constant value, which is unrelated to the amount of beads and the concentration of antigen.Optimal results of the colorimetric ELISA are achieved when the incubation volume meets each IMB＇s NEIV and is no longer enhanced by increasing the incubation volume.Thus, the reliable and relatively precise number of IMBs for ELISA detection during practical application could be determined.Most importantly, a study using IMB＇s NEIV would lay the foundation for a kinetics analysis of IMBs and antigens for future study.