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Sciex Developing Peptide Quantitation Assays Using Acoustic Ejection System


This story has been updated to correct the name of the Sciex technology from acoustic injection to acoustic ejection.

NEW YORK – Sciex has demonstrated the compatibility of its Echo MS acoustic liquid handling system with mass spectrometry-based peptide quantitation workflows, thus expanding its use beyond small molecule work to applications such as biopharma development and large-scale protein biomarker studies.

At the annual meeting of the US Human Proteome Organization in March, scientists from Framingham, Massachusetts-based Sciex, which is owned by life science giant Danaher, presented proof-of-principle data showing that the Echo MS system, which can inject up to three samples per second into a mass spec, can analyze peptides using a peptide quantitation workflow that makes use of stable isotope standards and capture by anti-peptide antibodies (SISCAPA).

The company is now beginning to develop peptide quantitation applications using the system, said Gavin Fischer, senior director of LC and integrated solutions at Sciex. While its initial target would likely be peptide quantitation within biopharmaceutical workflows, it could also extend to protein biomarker research, he added.

The peptides the company's researchers worked with in their method development included a set of 20 from the SARS-CoV-2 nucleocapsid protein, Fischer said.

Developed in collaboration with Sciex's Danaher stablemate Labcyte, the Echo MS system uses acoustical liquid handling for rapid injection of samples into a mass spectrometer. Sciex launched the system in 2020 and has since marketed it primarily for small molecule assays. The work detailed at US HUPO aimed to determine whether the system could also be used to analyze larger molecules like peptides and proteins.

The Echo MS could be of interest to protein biomarker researchers in that it would allow for extremely high-throughput experiments. For instance, Fischer noted that in the first project exploring use of the device for peptide measurements, the Sciex team quantified a pair of beta galactosidase peptides in a digest of the beta-gal protein containing around 100 peptides. With the Echo MS, they were able to quantify the two peptides in 10,260 samples in five hours. Had they done the same experiment using a conventional liquid chromatography mass spectrometry setup running a roughly seven-minute LC gradient, it would have taken almost 50 days of round-the-clock mass spec work.

"We were looking at very high-throughput peptide quantification," Fischer said, noting that the researchers set the system to inject at a rate of one sample every 1.5 seconds. "We got great reproducibility across the plate, great [coefficients of variation]."

The Sciex team followed this initial Echo MS work with experiments using a SISCAPA workflow, which uses upfront antibody enrichment of target peptides, to measure the 20 SARS-CoV-2 peptides. Christie Hunter, director of application and technical marketing at Sciex and leader of the effort, said in an email that SISCAPA was a promising technique for protein quantitation using the Echo because the upfront antibody enrichment simplified samples enough to where they could be analyzed without requiring LC separation.

"The key aspect here is finding the right upfront sample preparation approach that simplifies the proteomics sample, such that a flow injection approach is feasible," she noted. "The antibody enrichment of target peptides is very compelling as the right sample preparation workflow to pair with the Echo MS system because it is highly automatable, fast to perform, creates a highly enriched sample, and can be scaled based on sensitivity of target protein."

She added that such a workflow "could target labs that have a lot of proteomic samples to analyze and want to quantify a small target set of proteins."

Looking at limits of quantification for the 20 peptides, the researchers found that the system had an average LOQ of 1 femtomole per microliter with the most sensitive peptide assay, quantifying its target down to 0.4 femtomoles per microliter and at least to 3.7 femtomoles per microliter, Fischer said.

"The [US HUPO] presentation was a very solid demonstration that we could do things that were not small molecules," he said, adding that the company now plans to test "bigger and bigger molecules on the Echo system and on different mass specs and demonstrating the utility to customers as we expand into more markets with more workflows."

Asked whether it was realistic to think the system could be used for applications such as targeted quantitation of peptides in patient plasma, Fischer said that the company had not yet demonstrated this experimentally but that "there are theoretical reasons to believe that that will work."

He noted that Sciex has demonstrated the ability of the system to quantify small molecules in plasma.

"We are working on getting [the technique] into more and different matrices and exploring the upper size envelope, from peptides to moving our way up to whole proteins," he said. "We will continue to push the boundaries in all of those directions."

Sciex currently offers the Echo MS system coupled to its 6500 series mass spectrometers, but Fischer said that both among its outside customers and within the company, there is strong demand for combining the system with a time-of-flight instrument. He declined to confirm any specific plans for this, but indicated that the recently released ZenoTOF 7600 platform would be an obvious choice.

He said that perhaps the biggest challenge in combining the Echo MS system with a new mass spec — especially a TOF instrument — is on the software side.

"The problem with a TOF is, it generates a lot of data, and it does so very quickly," Fischer said. "And then if you speed up the sample delivery from once every couple of minutes to once a second, you start to have data transfer problems, data analysis problems. There are things you do in the background on the fly that you normally have several minutes to do that now you have several milliseconds to do."

He cited as one example the fact that he and his colleagues have had to upgrade the computer storage used for the experiments from hard drives to solid state drives because the hard drives were failing after a few weeks due to overuse.

Fischer indicated that the company plans initially to target the biopharma market with the system, noting that peptide quantitation is a key part of biopharma ADME workflows. Furthermore, "one can imagine that if you were to put the Echo in front of a TOF, you could move into the [biopharma] discovery space," he said. "A major driver for Sciex is getting into the biopharma market."

From there, the company will look to apply the technology to additional markets, including potentially protein biomarker research.

"We're very committed to the technology and we will continue to expand," Fischer said.

A similar system developed by AstraZeneca, Waters, and Labcyte actually predates the commercial launch of the Echo MS by about a year.

Waters has not taken any public steps to commercialize the system, and it is unclear whether it would be able to, given that Danaher owns both Labcyte and the Echo MS technology, but it is currently used internally at AstraZeneca, where scientists are developing a variety of applications on it, including high-throughput metabolomics screening and cell-based assays.