Additives for Diagnostics

What is Biolipidure®?

Biolipidure® is a BSA substitute for diagnostics additive.

Protein assays determine the amount of protein in an unknown solution. Specially, Western blotting is widely used for detection of protein which is a target protein or not. In Western blotting, prior to protein immobilization on the PVDF or nitrocellulose membranes, sample proteins are separated using SDS polyacrylamide gel electrophoresis providing information about molecular weight and the potential existence of different isoforms of the proteins using monoclonal or polyclonal antibodies. For sensitive detection of target proteins, non-specific binding of monoclonal or polyclonal antibodies on the membrane should be reduced.

NOF has supplied additives, Biolipidure®, for suppression of non-specific binding of antibodies on the membranes. By using of Biolipidure®, high sensitive Western blotting would be done.

Features

Biolipidure®, an additive for diagnostics, is non-biological origin synthetic compound and an alternative of BSA.

Suppression of non-specific adsorption

Enhancement of sensitivity and accuracy

Stabilization of antibody and enzyme

No lot-to-lot variations and No biohazardous risk

Additives for Diagnostics

Diagnostics are tools for detection of diseases and illnesses and for observation of health condition and are also useful for therapy and prevention. In general, blood, urine and stool are used as specimen for diagnostics. For reduction of suffering in patients and for sensitive detection of diseases, improvement of sensitivity in diagnostics has been required.

Biolipidure® has multiple functions such as suppression of non-specific adsorption of protein, stabilization of protein, and sensitization, all of which are required for developing diagnostic reagents. Biolipidure has variations with different molecular weights, hydrophilic-hydrophobic ratios, and ionic properties. You should test several to empirically determine which works best in your assay.

Blocking effect of Biolipidure®

Biolipidure® suppresses non-specific adsorption of biomolecule. NOF provides a set of products that can coat the surface of plastic, metal or other various materials. Biolipidure® consists of wholly synthesized polymers, so it is free from biohazardous risk and has little lot-to-lot variation, making Biolipidure® the best blocking agent for diagnostic reagents.

Blocking microplates

After a microplate was treated with Biolipidure®, the amount of non-specific adsorption of enzyme-labeled antibody was measured. The amount of non-specific adsorption of enzyme-labeled antibody was significantly reduced, proving that Biolipidure® provides excellent blocking effect.

Test procedure

200 µL of LIPIDURE® (diluted 10-fold by PBS) was applied to a 96-well plate.
The plate was aspirated and dried overnight.
100 µL of 24,000-fold diluted HRP-IgG was added.
After 1 hr incubation at a room temperature, the plate was washed with PBS-T.
TMB colorimetric substrate (KPL) was applied and the non-specific adsorption of HRP-IgG was measured.

Blocking magnetic beads

After magnetic beads was treated with Biolipidure®, the amount of non-specific adsorption of enzyme-labeled antibody was measured. The amount of non-specific adsorption of enzyme-labeled antibody was significantly reduced, showing that Biolipidure® provides excellent blocking effect.

Test procedure

170µg/mL of magnetic beads suspension was added to the same amount of Biolipidure® diluted 5-fold by TBS.
After 1 hr of incubation at a room temperature, the beads was washed with TBS.
10 ng of AP-IgG was added.
After 1 hr of incubation at a room temperature, the beads was washed with TBS–T.
Chemiluminescent substrate was applied and non-specific adsorption of AP-IgG was measured.

Blocking magnetic beads (Antibody solution)

The same protein-blocking effect was also achieved when Biolipidure® was added to an antibody solution. Biolipidure® was added to a diluted solution of enzyme-labeled antibody, and the amount of non-specific adsorption to magnetic beads was measured. The amount of non-specific adsorption of enzyme-labeled antibody was significantly reduced, proving that LIPIDURE® provides excellent blocking effect.
* Biolipidure® does not affect antibody titer.

Test procedure

0.1 times volume of Biolipidure® added to 100 ng/mL of AP-IgG solution.
100 µL of the AP-IgG solution containing Biolipidure® was added to 8.6 µg of magnetic beads.
After 1 hr of incubation at a room temperature, the beads was washed with TBS-T.
Chemiluminescent substrate was applied and non-specific adsorption of AP-IgG was measured.

Protein-stabilization effect of Biolipidure®

Biolipidure® stabilizes proteins, probably because the interaction between Biolipidure® and proteins maintains the native state structure of the proteins.

Stabilization of antibodies (Solid phase)

The stability of immobilized capture antibody was improved by treating the surface od the microplate by Biolipidure®. This treatment also reduces the non-specific adsorption of proteins applied subsequently.
After the capture-antibody-immobilized microplate was coated with Biolipidure® and stored at 25°C, the residual antibody titer was measured using ELISA (sandwich method).

By using Biolipidure®, the stability of antibody was improved, that is, alsmost 100% of antibody titer was maintained for more than 15 days.

Test procedure

Anti-mouse IgG antibody (capture antibody) was immobilized on a microplate.
After 1 hr of incubation at a room temperature, the plate was washed with PBS.
200 µL of LIPIDURE® diluted 10-fold by PBS was added.
After 1 hr of incubation at a room temperature, the plate was washed with PBS.
The microplate was stored at 25°C in desiccator.
Mouse IgG (antigen) was added.
After 1 hr of incubation at a room temperature, the plate was washed with PBS-T.
HRP labeled anti-mouse IgG antibody was added.
After 1 hr of incubation at a room temperature, the plate was washed with PBS-T.
Colorimetric substrate (TMB) was added and the antibody titer of the capture antibody was measured.

Stabilization of antibodies (Liquid phase)

LIPIDURE®-SF provides stabilization effect when added to a diluted solution of enzyme-labeled antibody. LIPIDURE®-SF was added to a diluted solution of enzyme-labeled antibody and residual enzyme activity was measured after storage at 4°C. By adding LIPIDURE®-SF, the stability of enzyme-labeled antibody was markedly improved and 100% of enzyme activity was maintained for 60 days.
* LIPIDURE®-SF does not affect antibody titer.

Test procedure

0.02 times volume of LIPIDURE®-SF08 was added to 10,000-fold diluted solution of enzyme-labeled antibody.
The solution was stored at 4°C.
The residual activity of labelled enzyme was measured with colorimetric substrate (TMB)

Sensitizing effect of Biolipidure®

BIOLIPIDURE® exhibits sensitizing effect in immunoassay systems such as latex agglutination or immunochromatography, most likely caused by the improved dispersion of latex particle or gold colloid.

Sensitizing effect of Biolipidure® in latex agglutination test

Biolipidure® provides excellent sensitizing effect in latex agglutination test.

Sensitizing effect of Biolipidure® in lateral flow test (immunochromatography)

Biolipidure® provides excellent sensitizing effect in lateral flow test (immunochromatography).

Suppression of lot-to-lot variation of Biolipidure®

For Suppression of Non-Specific Adsorption

Application examples of Biolipidure® ELISA / CLEIA

Biolipidure® provides sensitizing effect in sandwich type Chemiluminescent Enzyme Immunoassay (CLEIA)

Test proceedure

Mouse anti-Human IgG antibody was immobilized on a microplate.
After 1 hr of incubation at a room temperature, the plate was washed with TBS.
Biolipidure® solution (0.1 times volume) was added.
The plate was incubated for 1 hr at a room temperature.
Human IgG (antigen) standard was added.
After 1 hr of incubation at a room temperature, the plate was washed with TBS-T.
AP labeled anti-Human IgG antibody was added.
After 1 hr of incubation at a room temperature, the plate was washed with TBS-T.
Chemiluminescent substrate was added and calibration curve for Human IgG concentration was prepared.

Biolipidure®

Biolipidure-103 (10ml)

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Biolipidure-203 (10ml)

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Biolipidure-206 (10ml)

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Biolipidure-405 (10ml)

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Biolipidure-502 (10ml)

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Biolipidure-702 (10ml)