Tuesday, July 10, 2012

Did you know that BMG LABTECH’s software can do the ratiometric calculation necessary for the SoPRano™ Label-Free assay?

Fig.1: Brief Overview of SoPRano Assay™.
A protein ligand is added to the GNR-protein conjugate
and a spectrum is taken. The OD at λmax+80
is determined and is divided by
the new OD at λmax, to give the ratio
(ODλmax+80/ODλmax). Upon specific binding the
LSPR effect leads to a does dependent red shift in
λmax and thus an increase in the ratio of the OD values.
Working in conjunction with Pharma Diagnostics, BMG LABTECH has released a version of its MARS Data Analysis software (v2.40 or higher) which has the ability to easily and quickly perform the ratiometric calculations necessary for the SoPRano™ Label-Free assay.

The SoPRano™ label-free assay is based upon Localized Surface Plasmon Resonance (LSPR) which can be found in substances such as Gold Nano Rods (GNRs). GNRs have a specific absorbance maximum (λmax) and when proteins are subsequently bound to the GNRs, this λmax shifts to the red region of the spectrum. This means that different proteins and different protein concentrations bound to the GNRs will have a different change in λmax. Therefore the λmax and the change in λmax must be determined for each new SoPRano™ assay. To help with the data analysis, Pharma Diagnostics has created a ratiometric method that accurately measures this change in λmax.

Fig.2: Binding of hSA to its Antibody Using SoPRano™ LSPR.
Increasing amounts of the hSA antibody were added to GNRs
pre-conjugated with hSA. A dose-response binding curve
is produced using the ratiometric data
After determining the λmax of the SoPRano™ assay using fast, full-spectrum analysis, the λmax+80 is determined. This OD is then divided by the λmax OD to get ODλmax+80/ODλmax. Figure 1 shows graphically the λmax+80 changes that can be measured and Figure 2 shows the binding curves that can be created using the OD ratio. As pointed out earlier, these calculations are now easily done with BMG LABTECH’s MARS v2.40 or higher. In addition, a SoPRano™ MARS calculation template is available upon request. This template can be imported into MARS 2.40 and it will do all of the necessary calculations with one press of a button.

To learn more about the SoPRano™ assay please see our application note or contact your local BMG LABTECH representative.

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