Leveraging the core qPCR technology and validated assay library, MiRXES’ diagnostic arm has worked with numerous clinical and commercial partners to identify and validate biomarkers in both biofluids and tissues for a variety of diseases. The experience has enabled us to streamline and optimize a
highly controlled and automated workflow for high throughput quantification of 300-1000 human miRNAs from a minute amount of clinical specimen.

To date, more than 20,000 serum, plasma, urine and FFPE tissue specimens have been profiled using the workflow. MiRXES now offers this unique solution for projects that require greater data granularity and depth.

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1. Sample focus approach – this workflow employs a unique cDNA array approach, where hundreds of clinical sample cDNAs are loaded into the same 384-well PCR plate with intra-plate positive and negative samples for greater control and accuracy.

2. Absolute quantification – 6-log serial dilutions of synthetic miRNA templates are concurrently reverse transcribed and amplified with isolated sample RNA, enabling absolute quantification of miRNA target instead of relative Ct comparison.

3. Multi-layer control measures – six unique spike-in RNAs are added in high, medium and low concentration at distinct steps of the workflow to monitor and normalize isolation efficiency, RT-qPCR efficiency. Reference samples and RNAs are included as inter-study normalizers.

4. Advanced cDNA library – our unique approach to cDNA pre-amplification allows efficient and unbiased library preparation, thus enables the workflow to quantify up to 1000 miRNAs from RNA isolated from minute quantity of clinical samples (200 μl of biofluid, 2-3 standard FFPE tissue slices).

5. State-of-art bioinformatics – the use of multi-layer controls enables efficient data processing and normalization. Normalized data are analysed using multiple statistical methods including Principle Component Analysis, Data Randomization Test, Forward Logistic Regression and Support Vector Machine to identify potential biomarkers.

6. Diagnostic kit prototyping – upon validation of identified biomarker, diagnostic kit development services are available to translate the findings from bench to bedside.