The Application of NVoy Technology to Protein Stabilisation
A stable and reproducible protein standard curve is essential for applications such as enzyme production and
ELISA assays. Unfortunately some proteins have poor stability within the working range of the standard activity
assay. The addition of Stabil-P.A.C. NV10 to enzyme or protein standard stocks can stabilise standards and
contribute towards long term reliability, without compromising activity or protein structure.

PROTOCOL
Stability is very protein specific, but a general protocol is given below.

    1. Determine the protein concentration (using eg. Bradford assay, BCA assay, absorbance at 280nm).
    2. Typically a fivefold excess, by mass, of NV10 will protect the target protein. For example, use 200 μg/ml
        NV10 for 40 μg/ml protein.
    3. Each Stabil-P.A.C. tube contains 1.25 mg NV10 as a lyophilised powder.
    4. Add the protein solution to NV10 in Stabil-P.A.C. tubes to get the desired concentration, or make up a
        2.5 mg/ml stock of NV10 (1X stock) by adding 50 μL of buffer or distilled water to each Stabil-P.A.C. tube
        and add this stock to the protein solution.
    5. This protein / NV10 stock solution can now be used in subsequent dilutions to prepare sets of protein
        standards.
    6. NV10 1X stock solution can be stored for 1 week at 4oC or for longer term at -20 oC.

Troubleshooting
    • If the protein shows signs of aggregation or heavy losses the NV10 to protein concentration ratio can be
        increased, ie increase NV10 concentration and / or reduce protein concentration.
    • Alternatively, a lower NV10 to protein ratio can be used with proteins which have no history of
        aggregation.
    • Always measure assay blanks with buffer containing NV10.

EXAMPLE : Use of NV10 to Improve Enzyme Stability
The activity assay for citrate synthase is very sensitive, detecting protein in the μg/ml range. Standard activity
curves prepared using commercial citrate synthase require the stock protein to be highly diluted in 0.1 M Tris pH 7.8
in order to fall within the range of the assay. Unfortunately the resulting standard series is unstable, and activity is
quickly lost at the lower end of the assay. This requires the preparation of fresh standards for each experiment or
time point. The addition of NV10 to the citrate synthase stock before dilution results in a set of standards that is
stable for a prolonged time at 4 oC, saving time, maintaining reproducibility and reducing protein costs.

Summary
The addition of NV10 to unstable protein standards can result in a standard set which gives reliable and
reproducible results over several days.