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Suppl. Mater.: 

CLC number: R782.2

On-line Access: 2017-10-05

Received: 2016-08-11

Revision Accepted: 2017-02-19

Crosschecked: 2017-09-13

Cited: 0

Clicked: 4332

Citations:  Bibtex RefMan EndNote GB/T7714


Rui Wang


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Journal of Zhejiang University SCIENCE B 2017 Vol.18 No.10 P.845-853


Nominal effective immunoreaction volume of magnetic beads at single bead level

Author(s):  Rui Wang, Yuan Chen, Kai Fan, Feng Ji, Jian Wu, Yong-hua Yu

Affiliation(s):  College of Biosystems Engineering and Food Science, Zhejiang University, Hangzhou 310058, China; more

Corresponding email(s):   yhyu@zju.edu.cn

Key Words:  Nominal effective immunoreaction volume (NEIV), Immunomagnetic bead, Enzyme-linked immunosorbent assay (ELISA), Bead diameter, Mixing mode

Rui Wang, Yuan Chen, Kai Fan, Feng Ji, Jian Wu, Yong-hua Yu. Nominal effective immunoreaction volume of magnetic beads at single bead level[J]. Journal of Zhejiang University Science B, 2017, 18(10): 845-853.

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A1 - Rui Wang
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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.


方法:首先,采用两种直径的磁珠对50 µl不同浓度的抗原进行比色检测,证明磁珠上抗原的结合位点过量。随后,采用定量磁珠对相同浓度不同体积的抗原进行检测,得出"单个磁珠的有效免疫反应体积"这一概念。接着,采用三种不同用量的磁珠对浓度相同、体积梯度增大的抗原进行检测,以验证所提出的NEIV的概念。然后,通过测定孵育时长、孵育温度和震荡与否对比色结果的影响,找到对NEIV影响的关键因素。最后采用定量磁珠,对梯度稀释的抗原进行检测,建立磁珠用量与比色结果的关系,对NEIV的实际应用提出展望。


Darkslateblue:Affiliate; Royal Blue:Author; Turquoise:Article


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[22]List of electronic supplementary materials

[23]Fig. S1 Principle of EDC-NHS method for antibody modification of magnetic beads

[24]Fig. S2 Principle of IMB-based ELISA assay for PSA antigen detection

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