Posts Tagged ‘Bio-Rad Laboratories’

Droplet Digital PCR enables measurement of potential cancer survival biomarker

 :: Posted by American Biotechnologist on 01-09-2014

Researchers at Fred Hutchinson Cancer Research Center have used Droplet Digital PCR (ddPCR™) to demonstrate for the first time the quantification of a special class of tumor-attacking immune cell known to improve cancer survival, a subpopulation of T-cells called tumor-infiltrating T-lymphocytes or TILs. The study, led by Dr. Jason Bielas, Associate Member of the Public Health Sciences Division at Fred Hutch, paves the way for further study of the role of TIL quantification in immunotherapy and as a cancer survival predictor.

“Now that we have the sensitivity and ability to reproducibly count TILs in tumors, we may be able to stratify and more effectively treat patients based on tumor TIL count, especially with immunotherapeutics coming to market,” said Dr. Bielas, one of the lead authors of a paper reporting the TIL quantification results in Science Translational Medicine.

Quantifying TILs Using ddPCR

TILs directly attack tumor cells in a variety of cancer types. While the presence and quantity of TILs strongly correlate with increased patient survival, current tests are semiquantitative at best. As a result, TILs cannot be used for clinical decision making.

According to Dr. Bielas, TILs have a “genomic signature that can be digitally exploited.” This signature, which exhibits a vast amount of diversity, determines the genetic identity, or clonality, of the T-cell receptors (TCR) expressed on the surface of each TIL. With the advent of digital PCR – and the generation of tens of thousands of data points produced by Droplet Digital PCR – it is now possible to quantify these signatures, enabling the determination of the number of TILs.

“There’s no way you could do this with any method other than digital PCR because of the numerous primer pairs and probes that we have (45 forward primers, 13 reverse primers, and 30 probes),” said Dr. Bielas. “Digital PCR partitions all the reactions so you can amplify these targets independently of PCR efficiency without any competing side reactions.”

Fred Hutch researchers developed the Droplet Digital PCR-based “QuanTILfy” assay using Bio-Rad Laboratories’ QX100 ddPCR system. They then used QuanTILfy to count TILs, determine their frequency, and develop a grouping system to classify “clonality,” which might be a marker of druggable targets.

Study Results

Fred Hutch researchers performed the QuanTILfy assay on primary tumors from 30 ovarian carcinoma patients with known survival outcomes, ranging from 1 to 122 months. TIL frequency was approximately threefold higher in patients with a survival rate of more than five years compared with patients with survival rates of less than two years. These results show that higher TIL levels correlate positively with patient survival, consistent with the hypothesis that TILs play an active role in suppressing tumor formation.

The researchers also demonstrated that QuanTILfy can be used to accurately and reproducibly characterize T-cell clonality in patients with T-cell acute lymphoblastic leukemia. In each case, they saw a single QuanTILfy assay subgroup, indicative of clonal T-cell expansion. This finding was confirmed by deep sequencing.

The QuanTILfy assay proved to be both sensitive and accurate. In a mixture of human T-cells purified from blood and normal human lung fibroblasts, the assay demonstrated the ability to detect a single TCR rearrangement among 10,000 tumor cells. Importantly, it also demonstrated the ability of ddPCR technology to quantify a large number of markers simultaneously in a single reaction through multiplexing.

The Power of Partitioning

 :: Posted by American Biotechnologist on 12-30-2013

Bio-Rad Offers New PrimePCR™ Disease Panels for Real-Time PCR

 :: Posted by American Biotechnologist on 12-18-2013

Bio-Rad Laboratories announced the launch of more than 700 new PrimePCR human and mouse disease panels and 71 new pathway panels for real-time PCR (qPCR). The panels allow researchers to thoroughly and efficiently investigate genes known to be differentially expressed in a specified pathology or biological pathway. Bio-Rad now offers more than 1,100 panels with nearly 6,700 predesigned plate configurations, establishing Bio-Rad as the industry’s largest and most up-to-date provider of predesigned plates.

The expanded product line includes highly studied pathways such as Hedgehog and bone morphogenetic protein (BMP) signaling, as well as processes such as circadian rhythms and visual perception. In addition, disease categories span hundreds of conditions, ranging from specific cancers and neoplasms to metabolic diseases such as obesity and diabetes.

Working in collaboration with Thomson Reuters, Bio-Rad developed a plate design strategy that ensures that the most relevant gene targets are chosen for each real-time PCR panel; gene targets are weighted for differentially expressed genes and the frequency with which they appear in the peer-reviewed literature. For disease panels, Thomson Reuters referenced the diseases and their corresponding gene targets in the U.S. National Library of Medicine database.

All of the assays were designed strictly adhering to MIQE guidelines: maximum transcript coverage, minimal overlap with known single nucleotide polymorphisms, and spanning large introns where possible. In addition, all of the primer assays have been fully wet-lab validated for specificity, efficiency, and sensitivity to an unprecedented level of quality, according to Jan Hellemans, cofounder of the qPCR and bioinformatics firm Biogazelle, which worked with Bio-Rad to develop these assays.

Bio-Rad has also updated and expanded its existing predesigned pathway and collection panels to include the most scientifically relevant gene targets based on current literature and the latest bioinformatics data.

For more information on Bio-Rad’s PrimePCR products, please visit www.bio-rad.com/PrimePCR.

Labs for Inspiration: Making a Difference with Bio-Rad’s Science Ambassador Program

 :: Posted by American Biotechnologist on 11-20-2013

In August of 2012, Bio-Rad launched the Science Ambassador program, a corporate responsibility initiative fostering hands-on life science education. The program links interested scientists with interested nearby teachers through a simple website, so that the scientists can visit classrooms to conduct an exciting, one-hour DNA extraction lab. Bio-Rad facilitates these connections and supplies the scientists with free Genes in a Bottle™ DNA kits for up to 36 students per class, as well as easy-to-follow lesson plans.

In under a year’s time, the program has already made significant progress. Nearly 300 scientists and 300 teachers across the U.S. and Canada have signed up. And, in the single most important measure, more than 5,500 students (plus one big city mayor) have had the program’s signature lab experience: an up-close and personal encounter with their own DNA.

Read more…

What You Don’t See Truly Matters

 :: Posted by American Biotechnologist on 11-13-2013

All gels are not created equal when it comes to protein transfer. Mini-PROTEAN® TGX™ precast gels, together with the Trans-Blot® Turbo™ transfer system, offer you a complete solution for achieving fast electrophoresis and transfer times. In addition to speed, these innovative gels deliver maximum transfer of proteins onto membranes so you don’t see proteins left behind on the gel, ensuring quality western blotting results you can count on every time.

For more information visit Bio-Rad Laboratories TGX™ precast gel page.