High Load StrongZyme™ Streptavidin-(Poly)HRP
novel conjugate ideal for High Sensitivity detection in (pseudo)-1-step ELISA
tests with concentrated serum and plasma samples.
Recently some ELISA assemblers offer so-called pseudo-1-step assay test kits.
The distinguishing feature of these assay systems is a sequential addition of
all reagents with no wash between sample, biotinylated detector
antibody/antigen and Streptavidin-HRP (SA-HRP). Despite the tricky pipeting
associated with the risk of cross-contamination and imprecision these
systems are obviously accepted by certain customers in the blood donor
screening marketplace. Abandoning one or two washing steps
shortens assay time. Complementary advertised advantage is the smaller
consumption of wash buffer. Another, probably true, advantage is that human
serum/plasma matrix itself (as a rule applied in e.g. HBsAg and HIV-1/2 4G
test kits undiluted) works as an effective blocker reducing non-specific
binding of all detection system components. On the other hand false positives
or false negatives may occur through diverse "irrelevant circuits"
(essentially specific interactions apart of the assay components specificity)
associated with possible presence in the blood sample of antibodies against
biotinylated proteins, streptavidin and peroxidase.
Moreover, serum of the healthy adults contains from 0,1 to 0,4 nmol/L
of the endogenous biotin. Biotin weakens detection power of SA-HRP
through competing for the biotin-binding sites in conjugate that would
otherwise effectively react with the biotin label on the tracer antibody
and/or antigen. When using SA-PolyHRP conjugates inhibition through endogenous
biotin is much stronger compared to the conventional SA-HRP. The reason is
that in the SA-PolyHRP a molar SA/HRP ratio is heavily shifted towards HRP.
This means that the same working strength concentration of SA-PolyHRP
contains essentially smaller concentration of streptavidin and respectively
smaller concentration of the biotin-binding sites than conventional SA-HRP.
As a result SA-PolyHRP is basically less effective in pseudo-1-step ELISAs.
Otherwise presence of endogenous biotin in the clinical specimens necessitates
increase of the working strength conjugate concentrations to the levels that
make application of the SA-PolyHRP in pseudo-1-step ELISA test kits unlikely
Above problem is addressed now. Currently we offer novel High-Load
Streptavidin-HRP conjugate, item #HL-SZ-SA, that is especially designed for
the cost-effective applications in High Sensitivity ELISA test kits made in
pseudo-1-step format. Based on the advanced StrongZyme™ conjugation
chemistry #HL-SZ-SA has available an increased number of the biotin-binding
sites per mol of the bound HRP. Respectively, larger number of the
biotin-binding sites creates an extra "buffering" capacity against irrelevant
biotin present in the assay sample matrix making such conjugate less
sensitive to the endogenous biotin burden in serum and plasma specimens.