DOT ELISA AND

IMMUNOSTRIPS
DOT ELISA :
DOT ENZYME LINKED
IMMUNOSORBANT ASSAY
 The dot ELISA, a qualitative ELISA test,
can be performed more quickly with the
end detection done visually.
 Dot ELISA is a micro ELISA utilizing
antigen “dotted” onto nitrocellulose filter
discs that has been used for 25 years.
 Because of its relative speed and
simplicity, the dot ELISA is an attractive
alternative to standard ELISA
FABRICATION

 Polystyrene Chip
 Straight channels made via CNC
machining
 Surface modified via allydextran
 Thin film of PS between two PS sheets to
allow flow
 Nitrocellulose in well
LIMITATIONS

 Bubble Formation in Channels

 Non-specific binding
 Dot ELISA only qualitative
BUBBLES FORMATION IN
CHANNELS

 Caused by difference in fluid properties of

buffers
 Non-uniform distribution of reagents
 Causes faint lines in results
 Reduction in colormetric signal
NON-SPECIFIC BINDING

 Binding of secondary or non-target

molecules on
 reaction surface
 Low signal-to-noise ratio
 Better immobilization of monoclonal
antibody
 Surface modification
DOT ELISA

 DotELISA is only qualitative.

 Other ELISA methods, such as sandwich
ELISA
 would provide quantitative measurements
 Identification of novel clinical markers,
stricter environmental legislation, growing
consumer concern regarding food safety
and focus on disease self-management,
are just a few of the factors leading to the
requirement for an array of sensitive and
specific assays that are inexpensive,
easy to use and portable.
 with the development of a generic
quantitative immunostrip technology, that
may act as a platform for adaptation of
any laboratory based immunoasssay to a
quantitative rapid test format.
 The scope of application of the developed technology is
enormous. In recent years, advances in the production of
monoclonal antibodies has facilitated detection of
practically any desired analyte, from heavy metals to drugs,
and clinical markers to food pathogens using
immunoassay.
 The potential market for disposable, tests capable of rapid,
low cost, quantitative immunoassay is a billion dollar
market with applications in the sectors of clinical
diagnostics, food quality, police control, veterinary
diagnostics, military security and environmental monitoring,
to name a few.
 The team at BBG offers the development of an individually
tailored package, comprising reading device, sample
procurement and, if required, pre-treatment device and
disposable immunostrips. monitoring.
 Our technology:
 BBG possesses proprietary knowledge
in the area of molecular wiring, which
employs redox polymers with non-
diffusional mediators that can transfer
electrons from the redox centres of the
enzyme label (HRP/GOX) to the
electrode. Additionally, the group has
developed novel substrates for ALP
labels as well as the use of novel tailor
designed genetically modified enzymes
as labels, thus achieving very high
sensitivity (GOX = Glucose oxidase; ALP
= Alkaline phosphatase; HRP= Horse
radish peroxidase).
 BBG has developed immunostrips based on both
diffusional and non-diffusional transduction, capable of ng/L
(ppt) detection limits. Typical response times are 15-20
minutes and sample volumes can be as low as a few
hundred nanolitres. Typical immunostrip membranes are
employed and the electrodes are produced by thick film
technology, thus allowing a low cost production. The
immunostrip format can be modified for dual and multiple
analyte detection.
 The group has developed immunostrips for both
competition and sandwich immunoassays. Competition
assays are generally employed for small molecules and
sandwich for larger molecules.The team has also
established the biocompatibility of the immunostrip formats
with various solvents and possible sample matrices.
 IMMUNOSTRIP – SIMILAR TO
ELISA

Antibody 1 Antibody 2 Antibody 3

•monoclonal •monoclonal •specific to
antibody 1
•specific to antigen •specific to antigen
(one epitope) (second epitope) •fixed to
membrane
•tagged with colored •fixed to membrane
dye