K39 recombinant protein

K39 recombinant

Cat. Num. AAA505052

• Applications: ELISA, Western Blot, Dot Blot, ELISA, ELISA, Lateral Flow
• Synonyms: K39; K39 recombinant; RECOMBINANT ANTIGEN K39 from Leishmania chagasi; keratin; K39 recombinant protein

• Host: E Coli
• Form/Format: Dry Powder (Stabilized with 5% Trehalose)
• Concentration: 1.538 mg/ml (varies by lot)
• Sequence Length: 955

• Applicable Applications for K39 recombinant protein: ELISA (EIA); ; Possible Aplications:; Western Blot (WB), Dot Blot (DB), Indirect ELISA, Positive Control in Direct ELISA, CLIA, Lateral Flow (LF)
• Application Notes: Recommended concentration in ELISA plates: 2,2 ug/ml in plates for IgG detection; Suggested Titer by ELISA: approx. 1/11,850, which corresponds to 0.13 ug/ml of protein cncentration in plates for IgG detection.; Optimal dilutions/concentrations should be determined by the end user.; Other applications not yet tested, however should not be excluded.
• Recombinant Antigen: Lesihmania infantum K39 (Scalone et al., 2002).
• Note: Absence of precipitation after a freezing and thawing cycle is okay.
• Observation Notes: In some cases, purified proteins runat a molecular weight which is slightly different to thetheoretically calculated molecular weight maybe due to thehis-tag presence, which can produce a delay in SDS-PAGE.Proteins should be maintained frozen at high concentrations. The dilution to be performed for ELISA assays should be made with a small quantity of protein, thesame day of the experiment. In order to defrost the protein, maintain the aliquot at 25°C without shaking to avoid aggregation. Prior making test dilutions and after defros tthe protein, is recommended to remove possible protein aggregates by centrifuging the stock solution, avoiding alterations in the immobilization of the biomolecule to thesolid surface.
• Buffer: Before Lyophilization: 20 mM phosphate buffer pH 8, 1 M NaCl and 0.1% polyoxyethylene (10) tridecyl ether
• Batch Composition: Recombinant antigen with a his-tag in its N-terminus
• Reconstitution: With approximately 0.068 ml of sterile double-distilled water, a final concentration or approximately 1.538 mg/ml will be obtained. The solubilization of the cake should be developed for 15 minutes to allow a homogenous potein solution, considering that part of the cake can be on the glass-walls of the container. Upon reconstitution, leave the solution at least 15 minutes homogenizing with a mild agitation at 4°C. Avoid vigorous shaking that can cause foaming and protein denaturation.With this reconstitution, the protein will be maintained at pH 8. It is recommended that the users carry out their absorbance determinations to avoid concentration variabilities due to the equipment used, mainly in reproducibility analysis.
• Preparation and Storage: Lyophilized protein is shipped with on blue ice.; Upon arrival, it should be stored at 4°C to -20°C. Once reconstituted, it should be aliquoted in order to avoid repeated freezing and thawing cycles and store at 20°C to -80°C.

Protein concentration determined espectrophotometrically

(This protein does not contain any Trp residues. Experience shows that this could result in more than 10% error in the computed extinction coefficient Therefore, we have measured the protein concentration by using the colorimetric assay based on the interaction between Coomassie brilliant blue and the arginine and aromatic residues (Bradford Method) and its maximum absorption shifts from 470 nm to 595 nm. The standard curve was performed with the protein BSA. 40 μl of the protein were analysed.)

Purity control in SDS-PAGE: 15%

Titration curve by an ELISA assay

(In this plot, the optical density at 450/620 nm for positive (blue) and negative (gray) IgG sera are compared for each concentration of the recombinant antigen. An appropriate statistical test of significance for the comparison of means between both groups, the Welch's test, is employed. Eligible concentrations for the use of the antigen should present statistically significant differences between positive and negative sera. This happens when the intervals at the top do not overlap and, equivalently, when the p-value at the bottom is below 0.05. In the present figure, all p-values are below 0.05 and thus the intervals do not overlap. Therefore, any of the showed concentrations can be used to distinguish between positive and negative sera.)

Related Product Information for K39 recombinant protein

The Leishmania chagasi antigen k39-biotinylated has been prepared as a recombinant antigenfused to a his-tag. It is produced from the C-terminus (aa680-955) of the kinesin K39.

Product Categories/Family for K39 recombinant protein

PARASITES

References

Scalone, A., de Luna, R., Oliva, G., Baldi, L., Satta,G., Vesco, G., Mignone, W., Turilli, C., Mondesire,R.R., Simpson, D., Donoghue, A.R., Frank, G.R.,Gradoni, L. Evaluation of the Leishmania recombinantK39 antigen as a diagnostic marker for canineleishmaniasis and validation of a standardized enzymelinkedimmunosorbent assay. 2002, Vet. Parasitol.104:275-285.
Bradford, MM. A rapid and sensitive method for thequantitation of microgram quantities of protein utilizing theprinciple of protein dye binding. Anal Biochem. 1976,131:499-503.

NCBI and Uniprot Product Information

• NCBI GI #: 308885
• Molecular Weight: Determined by SDS-PAGE, the protein band is between molecular markers of 35,000 and25,000 Da, while relative molecular mass calculated fromamino acid sequence is 25,936.0 Da.
• NCBI Official Full Name: kinesin-like protein