Overcoming Protein Biomarker Identification Challenges

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Tamlyn Oliver: Hi, everyone. Welcome to Biocompare's Tech Insights Podcast, where we speak to scientific experts about new tools and technology that can help advance your research.

I'm Tamlyn Oliver, Managing Editor of Biocompare. Today's guest is Mike Mao, Senior Director of Research and Development at RayBiotech. He's going to talk to us about protein biomarker identification and antibody arrays as biomarker screening tools. Thanks for joining us today, Mike.

Mike Mao: Thanks for having me. I'm glad to be here.

Tamlyn Oliver: Awesome. We're going to be talking, like I said, about protein biomarkers and their identification, which can be challenging. So, we're hoping that in the time we have together, you can shed some light on the process as well as provide tips on how to overcome challenges. Are you ready?

Mike Mao: Sure.

Tamlyn Oliver: All right. Let's begin by having you explain some of these challenges associated with identifying protein biomarkers.

Mike Mao: Identifying protein biomarkers is rewarding but also challenging. The first challenge is the tremendous diversity of protein. Human genome consists of about 20,000 genes. With alternative splicing and posttranslational modifications, human proteome is estimated close to one million proteins. Finding the right protein biomarker is like finding the needle in the haystack.

Meanwhile, the wide dynamic range of these proteins in biological samples can expand quite a lot in concentrations, ranging from milligrams to femtograms per milliliter. Since clinically relevant biomarkers are often present in the sub-picogram to sub-nanogram per milliliter range, it requires highly sensitive tools to find them.

Tamlyn Oliver: Okay. How do you suggest that researchers overcome these problems to maximize their chances of biomarker discovery?

Mike Mao: ELISA, the enzyme-linked immunoassay, is the gold standard of protein quantification at such low concentrations. However, traditional ELISA detects only a single analyte at one time. It's time consuming and impractical to measures hundreds of analytes by ELISA because it requires such a large volume of samples.

Affinity-based antibody arrays are the ideal tools to biomarker discovery. They adopt the same detection methodology as ELISA while in high throughput reformat. In this instance, a large collection of distinct captured antibodies is mobilized onto a solid surface. The proteins of interest will be captured by these antibodies to achieve thorough analysis.

Analyte detection can be expanded by running multiple non-overlapping arrays in parallel to detect thousands of proteins in one experiment.

Tamlyn Oliver: All right. Let's talk about high density arrays next. What are some key factors that researchers should consider when planning to use them?

Mike Mao: There are two kinds of antibody arrays, sandwich-based or label-based. Sandwich-based arrays use two antibodies for each analyte. While paired antibodies increase the detection specificity, they also introduce complexity and possible cross reactivity. For this reason, a typical sandwich-based array panel can detect a few thousand analytes at most.

Label-based arrays, which are called the L-Series, use only one antibody fixed on solid surface with biotinylated samples. Since there are no cross-reactivity concerns among antibody, it allowed for the development of much higher density array panels. A common L-Series array can detect hundreds of analytes in each array.

When planning to use a high density array, several factors need to be considered; array format, sample volume, and data type. Sandwich-based arrays will have higher specificity, generally either quantitative or semi-quantitative data, but it will need a larger sample volume. Label-based arrays are semi-quantitative, but they require smaller sample volumes.

Tamlyn Oliver: Okay. So, what kind of sample material can be used on these high density arrays?

Mike Mao: Researchers have used our arrays to plot a diverse of sample types, including serum, plasma, cell culture supernatant, cell or tissue lyses, tears, saliva, milk, urine, cerebral spinal fluid, synovial fluid, and other body fluids.

Tamlyn Oliver: All right, a wide range of samples, clearly.

So, RayBiotech offers many different array products. What advice would you offer researchers to help them choose the right product for their particular study?

Mike Mao: There are a few considerations when planning a array experiment. First, how many analytes does the researcher need to detect? Second, does the research need quantitative or semi-quantitative data? Third, how much sample volume is available?

We have hundreds of premade arrays in stock in a wide range of panel sizes, from less than 10 to many thousands. Our array selection also covers 13 different species. If the researcher prefers to custom build their own analytes panel, they can choose from our library of 10,000 analytes.

Our largest array currently detects 6,000 human, 1,300 mouse, or 1,500 rat proteins for experiments. These are the L-Series arrays, which are semi-quantitative. Large panels are also available in the Quantibody line, where you can detect up to 1,200 proteins concurrently.

Whether quantitative or semi-quantitative, these high density rates are ideal for biomarker discovery. However, small arrays focused on a specific group of related proteins are very popular as well. If sample volume is not an issue for the researcher and only few dozen analytes are needed, our membrane-based arrays, which are called C-Series, are an excellent option that is compatible with any Western broad imaging system. In summary, RayBiotech can provide diverse products and service to meet your needs.

Tamlyn Oliver: Thanks, Mike, for sharing your protein biomarker identification expertise with us today. For more on protein biomarkers and the use of high density arrays for biomarker research, please read the related article, " Pushing the Limits of Multianalyte Immunodetection."

And thanks so much for listening. For more information on products, technologies, and the latest scientific advancements, please visit Biocompare.com, and have a great day.