{"id":4697,"date":"2019-05-13T18:44:00","date_gmt":"2019-05-13T18:44:00","guid":{"rendered":"https:\/\/www.waters.com\/blog\/what-do-great-vacations-and-peptide-bioanalysis-have-in-common-2\/"},"modified":"2022-07-28T03:12:23","modified_gmt":"2022-07-28T03:12:23","slug":"what-do-great-vacations-and-peptide-bioanalysis-have-in-common-2","status":"publish","type":"post","link":"https:\/\/www.waters.com\/blog\/what-do-great-vacations-and-peptide-bioanalysis-have-in-common-2\/","title":{"rendered":"What do great vacations and peptide bioanalysis have in common?"},"content":{"rendered":"

For starters no one wants a disastrous vacation, the same goes for a bioanalysis assay! What makes for a great vacation? A booking agent who understands you, on time transit, a room with a view a stones throw from the beach, friendly staff and great cuisine?<\/p>\n

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Knowing that you are in safe hands and leaving nothing to chance each and every step of the way goes a long way to peace of mind!<\/p>\n

Choosing a trusted travel agent is a bit like investing in a complete peptide and protein bioanalysis solution.<\/p>\n

Often bioanalysts evaluate the sensitivity, accuracy, and precision of data from a high performance triple quadrupole mass spectrometer as a marker of assay performance, but how often is the influence of good sample handling, preparation, and separation overlooked?<\/p><\/blockquote>\n

As mentioned in part 1\u00a0of this blog series “Where Did My Sample Go?<\/a>“, ignoring non-specific binding can be catastrophic for sample recovery of peptides and proteins, the worst case scenario being complete sample loss. Who wants to lose their sample before they even inject? In part 2 of the series “Lost Samples in the Container: Non-specific Binding and the Impact of Blocking Agents<\/a>“, Moon highlighted that traditional methods of mitigating non-specific binding (addition of carrier protein, blocking agents, and modification of solution conditions\u2026) are generally not a one size fits all approach and less than ideal.<\/p>\n

That\u2019s why we have developed QuanRecovery Vials and Plates with MaxPeak High Performance Surfaces (HPS),\u00a0to be compatible with downstream LC-MS. MaxPeak HPS are new and innovative technologies designed to increase analyte recovery, sensitivity, and reproducibility by minimizing analyte\/surface interactions that can lead to sample losses.<\/p>\n

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Figure 1. % Recovery of pramlintide using QuanRecovery with MaxPeak (HPS) or standard polypropylene.<\/em><\/figcaption><\/figure>\n

In Caitlin Dunning’s application note\u00a0“Development of a SPE LC-MS\/MS Method Utilizing QuanRecovery Sample Plates with MaxPeak High Performance Surfaces for the Bioanalytical Quantification of Pramlintide from Serum<\/a>” we show the impact of using a standard polypropylene plate on sample recovery for analysis of a challenging hydrophobic peptide: pramlintide (Figure 1).<\/p>\n

So that\u2019s sample handling, but what about sample preparation? That peptide of interest doesn\u2019t come on its own. We often need a highly selective sample preparation method that can isolate the target from other related species in the sample matrix. But where do we start?<\/p>\n

That\u2019s where the\u00a0Oasis \u00b5Elution Peptide Separation Technology (PST) Method Development Plate<\/a> comes in handy to optimize your sample preparation method for maximum selectivity and recovery. Usually, a mixed-mode solid-phase extraction (SPE) sorbent, from the\u00a0Oasis SPE family of sorbents,\u00a0is the best way to clean-up and concentrate the sample (Figure 2). They employ both an ion exchange and a reversed-phase retention mechanism, which provide orthogonal modes of analyte isolation and cleanup. Brilliant!<\/p>\n

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Figure 2. A<\/strong>. Screening multiple SPE sorbents (Oasis MAX and WCX) using Waters \u00b5Elution PST Method Development Plate to quickly assess recovery and matrix effects for pramlintide. B<\/strong>. With an optimized protocol the recovery of pramlintide was increased to 75%.<\/em><\/figcaption><\/figure>\n

The Oasis 96 well-plate enables samples to be eluted in a small volume, eliminating the need for dry down and reconstitution, but as Kim reminded us in part 3\u00a0of this blog series, “Sample Prep before LC-MS Quantification of Peptides and Proteins<\/a>“, samples can still be lost during storage in the autosampler. Now arriving,\u00a0QuanRecovery 96 well-plates with MaxPeak HPS<\/a>!<\/strong><\/p>\n

So hopefully you\u2019re convinced handling and preparation of samples is important. What about LC separation? Is your peptide polar, hydrophobic, or on the larger size? Matching your peptide\u2019s physicochemical properties to the right column from\u00a0our range of peptide bioanalysis column technologies\u00a0can ensure sharp peaks and optimal separations (Figure 3).<\/p>\n

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Figure 3. Column Chemistry for Peptide Separations.<\/em><\/figcaption><\/figure>\n

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In Caitlin’s application note, to get the very best quantitative performance using class leading\u00a0ACQUITY UPLC I-Class PLUS and\u00a0Xevo TQ-XS\u00a0LC-MS technology, it was important to use our complete solution to achieve ultimate sensitivity, accuracy and precision:<\/p>\n