IMMUNOASSAY UTILIZING MAGNETIC BEADS FOR RAPIDVIRUS DETECTION IN THE MICROFLUIDIC FLOW CYTOMETER SYSTEM

Sung-Yi Yang, Kang-Yi Lien, Kao-Jean Huang, Huan-Yao Lei, Gwo-Bin Lee

National Cheng Kung University, Tainan Taiwan

uTAS2007

Professor : Liu, Cheng-Hsien   Student : Olof Sterner

AGENDA

1. Introduction2. Design and Fabrication3. Results4. Conclusion5. Discussion

In this article: Integrated fluid chip for rapid

virus identification. Fluid movement is performed by

micro circulatory pump driven by airpressure.

Magnetic beads covered with specific antibodies.

Detection of virus is made through fluorescence by a PMT module.

INTRODUCTION

Current viruses threatening mankind Influenza Hepatitis HIV Dengue fever Ebola Yellow Fever Etc

Treatment for viral infections are far behind those of bacterial infections Quick diagnosis crucial

INTRODUCTION

DESIGN AND FABRICATION

Magnetic beads Covered in specific

antibodies toward a certain virus

Virus particle functions as antigen and binds specifically to the primary antibody on the magnetic bead.

Secondary antibody towards the same virus but targeting a different epitope binds to the virus and carries a fluorescent compound.

DESIGN AND FABRICATION

Main detection principle1. Magnetic beads with

primary target antibody

2. Target antigen is added (Virus)

3. Secondary fluorescent antibodies is added

4. Fluidic focusing

5. Detection by means of fluorescense

DESIGN AND FABRICATION

The integrated flow cytometer system

A

B

C

Dimensions : 46 mm * 66 mm * 5 mm

DESIGN AND FABRICATION

A – Virus attachement to magnetic beads.

1. Injection of bio sample and magnetic beads

2. Introduction of sec fluorescent antibodies

3. Mixing with rotary microfluidic pump

DESIGN AND FABRICATION

Airpressure driven rotary microfluidic pump

Biomed Microdevices (2007) 9:545–554DOI 10.1007/s10544-007-9062-6

DESIGN AND FABRICATION

B – Washing and Pretreatment

1. Cu microcoils induced magnetic field attracts magnetic beads

2. Washing with buffer3. Release of magnetic beads(Flow generated by rotary microfluidic pump)

Purification and enrichment of virus samples utilizing magnetic beads on a microfluidic system, Lab Chip, DOI: 10.1039/b700516d, (2007)

DESIGN AND FABRICATION

C – Fluid focusing, detection and sorting

1. Sheath flow injection2. Flow from rotary microfluidic pump for fluid

focusing3. Detection by PMT module4. Sorting of magnetic beads using feedback

signals from PMT5. Collection of magnetic beads(Flow generated by rotary microfluidic pump

RESULTS

Evaluation of:

Pumping rate vs driving frequency of circulatory pump

Magnetic field and Temperatur vs Applied current

Fluorescence signal amplitude vs timeVirus detection limit

RESULTS

Pumping rate vs driving frequence of circulatory pump

Magnetic field and Temperature in focusing vs Applied current

RESULTS

Signal amplitude from fluorescence measurement vs time

Minimum concentration of 104 pfu/ml of Dengue virus could be detected.

RT-PCR has detection limit of ~1 pfu/ml

Singal to Noise ratio of 30

CONCLUSION

Design for rapid virus

diagnosis has been proposed

DISCUSSION

Easy principle, sandwich immunoassay is well documented

Gate design is not included in article

Sensitivity needs to be enchanced

REFERENCES

Lab Chip, 2007, 7, 868–875, DOI: 10.1039/b700516d

Biomed Microdevices (2007) 9:545–554 DOI 10.1007/s10544-007-9062-6

TACK!