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What is Liquid biopsy?

Liquid biopsy is a non-invasive diagnostic technique that allows the detection of tumor material by a simple peripheral blood sample. This technique will serve to improve the diagnosis, prognosis and follow-up of any tumor, and estimate its probabilities of metastasis, the event with the greatest clinical impact today in oncology. Liquid biopsy is considered the technology with the greatest potential for impact on the oncology diagnostic market.

Nasasbiotech is developing new approach in the field of liquid biopsy from the analysis of circulating tumor cells (CTC), the circulating tumor DNA (ctDNA) and the extracellular vesicles of the peripheral blood of the patient, which contain proteins, RNA and DNA with which they mediate cellular communication, with special impact on the metastasis process.

The competitive advantage of extracellular vesicles as biomarkers is based mainly on their greater stability and integrity and on their relative abundance associated with the process of tumor progression.

What is ExoGAG?

ExoGAG is a patented exosome purification method that allows almost all exosomes to be isolated from a liquid biopsy sample with a minimum amount of co-precipitated material, such as protein (mass spectrometry analisys, western-blot, elisa, protein arrays, etc) or genetic material (ADN, ARN or micro RNA exosome analisys as PCR, digital PCR, Beeming, secuenciation, mutation analisys, etc.), which makes it an ideal product for the search of biomarkers.

PROTOCOL

EXO-GAG SERUM/PLASMA/URINE EXOSOMES ISOLATION PROTOCOL

1. Collect the plasma/serum/urine sample. Samples can be frozen until the moment in which we want to use them; if the sample has been frozen, thaw it and temper it before processing). 2. Centrifuge the sample at 2000 x g for 10' to remove cells and cell debris. 3. Transfer the supernatant to a new tube and discard the pellet of possible cell debris.

PROTOCOL

EXO-GAG SERUM/PLASMA/URINE EXOSOMES ISOLATION PROTOCOL

4. Add the volume of sample to isolate exosomes to a new tube and add twice the volume of reagent A. That is, use a sample/reagent A ratio 1:2; (for example, to process 500 ul of plasma, add 1000 ul of precipitation reagent). 5. Mix the sample and precipitation reagent by inverting the tube or vortexing to homogenize the final solution (the solution will have a characteristic blue color). 6. Incubate the sample for 5 'at 4oC.

PROTOCOL

EXO-GAG SERUM/PLASMA/URINE EXOSOMES ISOLATION PROTOCOL

7. Centrifuge the sample at 15,800 x g; 15 'at 4oC. 8. Remove the supernatant being careful not to remove the pellet containing the exosomes (this pellet will be dark blue). 9. Resuspend the exosomes in the appropiate buffer, depending on the technique and analysis, as protein analisys (mass spectrometry analisys, western-blot, elisa, protein arrays, etc) or genetic material analisys (ADN, ARN or micro RNA exosome analisys as PCR, digital PCR, Beeming, secuenciation, mutation analisys, etc.)