antibody which is known as secondary antibody immobilized at control line.
Further steps involved in Sandwich ICA are as follows:
• At the beginning, analyte is applied to the sample pad where it migrates through
conjugate pad.
• Then the analyte present at the conjugate pad is captured by labelled antibody
and an analyte-antibody complex is formed.
• Next, the complex migrates towards nitrocellulose membrane where it binds
with primary antibody present on the test line and thus a sandwich is formed.
• Remaining labelled antibody conjugate will be bounded by secondary antibody
present on the control line.
Occurrence of test line confirms presence of the analyte and the color intensity of
the line determines the concentration of the analyte. A portable strip reader can
quantitatively measure this intensity.
2.2 Competitive ICA
It is also known as inhibition immunoassays. These kinds of assays are mostly
suitable for detection of low molecular weight compounds but can also be applied
to large molecular weight compounds. Steps involved in competitive ICA are:
• First the analyte to be detected is applied on the sample pad which will be then
migrated to the conjugate pad.
• Then the prefixed antibody present at the conjugated pad will be hydrated and
start flowing with the moving sample.
• After that the sample reaches at the nitrocellulose membrane and migrate
through the membrane towards the zone of control and test lines. The test line
contains pre-immobilized antigen which will bind specifically to labelled conjugate and control line contain pre-immobilized secondary antibody which also
have the affinity for labelled conjugate. If the target analyte is present in sample
solution, there will be a competition between the antigen to be detected and the
one which is already immobilized at test line of strip to bind with labelled
conjugate [24]. However, in absence of any analyte, labelled conjugate will
form a complex with the secondary antibody present at the control line and will
show visual line at the control line only as shown in Fig. 4.
2.3 Multiplex ICA
This type of immunoassay is mainly used to detect multiple analytes present in the
sample. Multiple test lines appear in this type of format as shown in Figs. 5 and 6.
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M. Pant et al.
Further steps involved in Sandwich ICA are as follows:
• At the beginning, analyte is applied to the sample pad where it migrates through
conjugate pad.
• Then the analyte present at the conjugate pad is captured by labelled antibody
and an analyte-antibody complex is formed.
• Next, the complex migrates towards nitrocellulose membrane where it binds
with primary antibody present on the test line and thus a sandwich is formed.
• Remaining labelled antibody conjugate will be bounded by secondary antibody
present on the control line.
Occurrence of test line confirms presence of the analyte and the color intensity of
the line determines the concentration of the analyte. A portable strip reader can
quantitatively measure this intensity.
2.2 Competitive ICA
It is also known as inhibition immunoassays. These kinds of assays are mostly
suitable for detection of low molecular weight compounds but can also be applied
to large molecular weight compounds. Steps involved in competitive ICA are:
• First the analyte to be detected is applied on the sample pad which will be then
migrated to the conjugate pad.
• Then the prefixed antibody present at the conjugated pad will be hydrated and
start flowing with the moving sample.
• After that the sample reaches at the nitrocellulose membrane and migrate
through the membrane towards the zone of control and test lines. The test line
contains pre-immobilized antigen which will bind specifically to labelled conjugate and control line contain pre-immobilized secondary antibody which also
have the affinity for labelled conjugate. If the target analyte is present in sample
solution, there will be a competition between the antigen to be detected and the
one which is already immobilized at test line of strip to bind with labelled
conjugate [24]. However, in absence of any analyte, labelled conjugate will
form a complex with the secondary antibody present at the control line and will
show visual line at the control line only as shown in Fig. 4.
2.3 Multiplex ICA
This type of immunoassay is mainly used to detect multiple analytes present in the
sample. Multiple test lines appear in this type of format as shown in Figs. 5 and 6.
238
M. Pant et al.