Agilent and BioTek Collaborate on Seahorse XF Normalization
The power of the well-known Seahorse XF technology from Agilent Technologies lies in its ability to simultaneously and kinetically measure mitochondrial respiration and glycolysis – two major energy pathways – in live cells. By doing so, researchers can closely monitor these dynamic events in real time rather than using separate assays for each cellular activity. As mitochondrial respiratory rates correlate to cell number, small differences in the number of adherent cells present in each microplate well create differences in the data gleaned from the Seahorse XF workflow. Normalizing XF data allows researchers to compare experimental data from well-to-well, treatment-to-treatment, plate-to-plate and more.
Previously, Agilent provided guidance on the range of normalization strategies allowing customers to develop and validate their own normalization protocols, a time-consuming process. Mr. Ned Jastromb, Product Manager for New Platforms in the Cell Analysis Division at Agilent, and others on the Seahorse XF team noticed that researchers were erroneously publishing information based on raw data, not normalized data. In fact, in over 4000 published papers using Seahorse XF technology, less than half incorporated normalized data. “In order for Agilent to continue to set the global standard for measuring metabolic function, we needed to offer an easy, standardized way for our customers to normalize and report data,” recalls Mr. Jastromb. At the same time, he wanted a way for researchers to self-evaluate any potential sample loss or other anomalies in the rigorous cell culture workflow.
When evaluating potential options to achieve the two goals, Mr. Jastromb ultimately turned to BioTek Instruments. “We wanted to partner with the leader in imaging that could deliver on a cost-effective, automated and validated solution,” he notes. “BioTek and their imaging products have a great reputation in this space, and by connecting our respective technologies, we knew we could create a synergistic experience for researchers.” Together, both sides quickly aligned and created a detailed plan for a validated solution that linked Agilent’s Seahorse XF technology with BioTek’s Cytation™ Cell Imaging Multi-Mode Readers.
In the integrated and optimized system, researchers capture a brightfield image of their cells using BioTek’s Cytation prior to metabolic analysis on the Seahorse XF analyzer. The image provides visual feedback and an opportunity to quality control the cell seeding conditions; any errant wells may be excluded from analysis. This improves live cell assay reproducibility and data quality. After analysis, cell count-based normalization is performed using fluorescence images captured with Cytation which opens up the potential for true experimental comparison. Cytation and the Seahorse XF analyzer are both integrated with a mutual control interface so that they seamlessly talk to each other and enhance ease-of-use. These images then get transferred to the XF results file so researchers can use this documentation procedure to improve their XF assay success rates.
“We always felt that we were in good hands with BioTek,” says Mr. Jastromb. “What we didn’t realize at the onset was just how receptive customers were to the value and flexibility provided by Cytation.” Customers find the system easy to learn and use, and the simplified procedure doesn’t create disruption in the overall assay workflow. Additionally, they love being able to associate the brightfield image with their Seahorse XF experiments and by doing so enhance their biologically meaningful conclusions. Beyond supporting the measurement of metabolic functions, Cytation also enables a broad range of additional cell-based research opportunities from basic to advanced.
Both companies are actively promoting the combined solution, including educating customers through workshops, conferences, lunch & learn seminars and more. The application note titled, “XF Data Normalization by the Agilent Seahorse XF Imaging and Normalization System” details the normalization protocol using the combined system. Mr. Jastromb summarizes the Agilent-BioTek partnership by saying, “I look forward to collaborating with BioTek on new projects as together we can open the door to future possibilities that ultimately benefit researchers.”
To learn more about Agilent Technologies, visit their web site.
Thanks to Ned Jastromb at Agilent Technologies for sharing his BioTek experience.