Report
Teaser, summary, work performed and final results
Periodic Reporting for period 1 - MULTIDET (Multiplex qPCR detection and chemical analysis towards multifunctional diagnosis)
Teaser
Chronic kidney disease (CKD) affects 10% of the population worldwide and sporadic cases of acute nephritis, if not detected early enough, may also progress to a chronic form. In particular, glomerulonephritis (GN) represents the main cause of chronic renal failure (25%). GN...
Summary
Chronic kidney disease (CKD) affects 10% of the population worldwide and sporadic cases of acute nephritis, if not detected early enough, may also progress to a chronic form. In particular, glomerulonephritis (GN) represents the main cause of chronic renal failure (25%). GN comprises several kidney diseases and its presentation depends on the specific pathology, therefore, rapid and accurate diagnosis is crucial because the outcome and the treatment differ in different diseases.
Despite the existence of several cheap traditional tests, based on the analysis of urinal and blood samples, there is no technology currently available to measure simultaneously DNA biomarkers, associated with infective GN, and to perform integrated cellular analysis.
The development of fast multiplex quantitative PCR (qPCR) technologies offers a unique opportunity to propose a Point of Care Test (PoCT) able to perform new multifunctional cheap diagnostics.
MULTIDET project aims at developing and optimizing multiplex ultrafast qPCR for the diagnosis of bacterial GN from urine samples, determining microorganisms’ identity and pattern of resistance/susceptibility to antibiotics. Moreover, the possibility to perform an integrated analysis of cellular biomarkers will allow the differential diagnosis of infective and non-infective GN.
Work performed
The main research and development project of ELVESYS is FASTGENE, the technology at the heart of MULTIDET project. FASTGENE is able to carry out ultrafast thermal cycling of the sample located in a patented microscopic optofluidic chip whose temperature is controlled by a micro thermal exchanger.
In order to process biological specimens and perform analysis of their molecular and cellular contents, a prototype processing bacterial samples and performing ultrafast real-time qPCR has been designed and developed and a relative patent was filed. In order to increase the sensitivity of microorganisms’ detection, PCR conducted in gel phase has been optimized and integrated. PCR in gel phase leads to the formation of so called, molecular or polymerase colonies (polonies), where each polony is the result of the amplification of a single molecule of target DNA or RNA. This is possible because the gel limits the diffusion of nucleic acids and the amplification products remain closed to the parental molecule, forming a detectable polony.
Therefore, this technology offer the advantage of directly quantifying the amount of target genetic material present in the sample, by simply counting the number of polonies formed in the gel. Additionally, the spatial separation of polonies reduces the competition between targets and consequent mismatches, as the genetic material remains confined in a restricted space of the gel, allowing high multiplex analysis and remarkably increasing the sensitivity of molecular diagnostics conducted with this technique.
Finally, a market research was conducted, in order to identify potential customers and competitors and define market’s needs in the context of Point of Care Tests (PoCT) and, specifically, of kidney diseases diagnosis.
Final results
With FASTGENE technology, ELVESYS is proposing a valuable alternative to the tests currently available for the diagnosis of kidney diseases. Diagnostic methods commonly utilized are based on the measurement of proteinuria, and albumin and creatinin urinary concentrations. Additional information can be provided by the presence of blood in the urine and through the measurement of blood electrolytes and blood urea nitrogen. Nevertheless, these tests are time consuming and often unreliable, requiring additional invasive confirmation analysis. Moreover, as kidney diseases comprise several different pathologies, their rapid and precise identification is of great importance for the outcome of the disease.
FASTGENE solution will be able to combine ultrafast multiplex qPCR for the characterization of bacterial glomerulonephritis with the analysis of cellular biomarkers, using an integrated and completely automatic plug & play system for the rapid and simultaneous differential diagnosis of infective and non-infective kidney diseases, minimizing the amount of starting material required and thus the costs.
The application of this technology will provide a faster and effective solution for a group of pathologies having a great impact on the global health, granting a rapid and personalized therapy able to reduce chronic renal failure events and antibiotics resistance spreading.
Moreover, it will benefit the management of microbiology lab, by reducing the analysis and the personnel required and thus the overall costs.