TY - JOUR
T1 - Paper-based Vertical Flow Immunoassay (VFI) for detection of bio-threat pathogens
AU - Chen, Peng
AU - Gates-Hollingsworth, Marcellene
AU - Pandit, Sujata
AU - Park, Anson
AU - Montgomery, Douglas
AU - AuCoin, David
AU - Gu, Jian
AU - Zenhausern, Frederic
N1 - Publisher Copyright:
© 2018 The Authors
PY - 2019/1/1
Y1 - 2019/1/1
N2 - Currently, the standard method for identifying biological agents of potential threats to national security and public health, such as pathogens, virus, and toxins, mainly rely on microbiological cultivation. This method is time-consuming and it requires sophisticated equipment and well-trained personnel, which are often unavailable in remote areas or at point-of-need. Therefore, an alternative rapid, simple, and sensitive method for detecting bio-threat agents is in crucial need. We report a paper-based Vertical Flow Immunoassay (VFI) device that can overcome these limitations. The VFI device utilizes a nanoporous nitrocellulose membrane encapsulated in a stainless steel filter holder. As the sample is pushed through the membrane, which is pre-functionalized with capture antibody, a sandwich assay is formed and colorimetric signal is generated to reflect the presence of target antigens. Through theoretical analyses of antigen-antibody binding process inside a porous membrane, we identified two critical factors – membrane pore size and sample flow rate that can be optimized to improve the assay sensitivity. Then, the effects were demonstrated through experimental studies using Burkholderia pseudomallei (the causative agent of melioidosis) as a model pathogen. The B. pseudomallei VFI was based on an immunoassay targeting the B. pseudomallei surface capsular polysaccharide (CPS). The experimental results agreed well with the theory showing that increasing the flow speed (up to 1.06 mm/s) and reducing the membrane pore size (down to 0.1 µm) could improve the sensitivity by at least 5 times. The VFI's limit-of-detection for CPS spiked in buffer solution was determined to be 0.02 ng/mL. The developed VFI shows great potential as a point-of-care tool for detection of bio-threat agents in a variety of clinical and resource-restricted conditions.
AB - Currently, the standard method for identifying biological agents of potential threats to national security and public health, such as pathogens, virus, and toxins, mainly rely on microbiological cultivation. This method is time-consuming and it requires sophisticated equipment and well-trained personnel, which are often unavailable in remote areas or at point-of-need. Therefore, an alternative rapid, simple, and sensitive method for detecting bio-threat agents is in crucial need. We report a paper-based Vertical Flow Immunoassay (VFI) device that can overcome these limitations. The VFI device utilizes a nanoporous nitrocellulose membrane encapsulated in a stainless steel filter holder. As the sample is pushed through the membrane, which is pre-functionalized with capture antibody, a sandwich assay is formed and colorimetric signal is generated to reflect the presence of target antigens. Through theoretical analyses of antigen-antibody binding process inside a porous membrane, we identified two critical factors – membrane pore size and sample flow rate that can be optimized to improve the assay sensitivity. Then, the effects were demonstrated through experimental studies using Burkholderia pseudomallei (the causative agent of melioidosis) as a model pathogen. The B. pseudomallei VFI was based on an immunoassay targeting the B. pseudomallei surface capsular polysaccharide (CPS). The experimental results agreed well with the theory showing that increasing the flow speed (up to 1.06 mm/s) and reducing the membrane pore size (down to 0.1 µm) could improve the sensitivity by at least 5 times. The VFI's limit-of-detection for CPS spiked in buffer solution was determined to be 0.02 ng/mL. The developed VFI shows great potential as a point-of-care tool for detection of bio-threat agents in a variety of clinical and resource-restricted conditions.
KW - Biological threat reduction
KW - Burkholderia pseudomallei
KW - Melioidosis
KW - Paper-based microfluidic
KW - Point-of-care diagnostic
KW - Vertical Flow Immunoassay
UR - http://www.scopus.com/inward/record.url?scp=85051966452&partnerID=8YFLogxK
UR - http://www.scopus.com/inward/citedby.url?scp=85051966452&partnerID=8YFLogxK
U2 - 10.1016/j.talanta.2018.08.043
DO - 10.1016/j.talanta.2018.08.043
M3 - Article
C2 - 30262102
AN - SCOPUS:85051966452
SN - 0039-9140
VL - 191
SP - 81
EP - 88
JO - Talanta
JF - Talanta
ER -