Biomarkers of animal health and welfare
The activity will consist in 3 subprojects:
- Refinement of the time resolved fluorescent immunoassays (TRFIAs) for chromogranin, alpha-amylase and amyloid-A in saliva, and adaptation to other veterinary species and humans.
- Development of automated immunoturbidimetric assays for total adiponectin and high-molecular weight (HMW)adiponectin measurements in dogs and if possible in other species.
- Development and clinical validation of a real-time PCR technique for detection of Leishmania infantum and of a sensitive assay for quantification of anti-leihmania antibodies in canine saliva.
Basically we will refine and optimize our previously developed assays by various technical improvements in order to increase sensitivity, reduce time of assay and volume of reagents. Also these assays will be adapted and validated for saliva measurements in other veterinary species such as horses, sheeps and dogs and also in humans.
The automated immunoturbidimetric assays have several advantages since they allow to increase the sample throughput as well to reduce the imprecision due to manual pipetting. Also they avoid the need of accumulate samples, as is the case of the traditional ELISA tests, since single samples can be measured without waste of reagents.
In this project we will develop automated immununoturbidimetric assaya for adiponectin and HMW-adiponectin in dogs. Also it will be explored if this assay can have cross-reactivity with other animal species and even if it could be adapted to humans. Since despite the importance of these proteins in obesity and other metabolic and inflammatory diseases there are no automated assays developed for its quantification in any veterinary species; and both proteins are in enough high concentrations to be quantified by immunoturbidimetry.
In this project we will develop a real-time PCR fluorometric technique that will allow for first time the quantification of leishmania in saliva. In addition we will develop a sensitive fluorometric assay to quantify antibodies against leishmania in saliva and we will compare it with the conventional techniques used in serum.