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bioRxiv · 10.1101/2020.03.13.990192

Detection of M-Protein in Acetonitrile Precipitates of Serum using MALDI-TOF Mass Spectrometry

Mehra, N.·Gopisetty, G.·Subramani, J.·Rajamanickam, A.·Sundersingh, S.·Karunakaran, P.·Perumal Kalaiyarasi, J.·Kannan, K.·Radhakrishnan, V.·Tenali Gnana, S.·Thangarajan, R.
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Abstract

Purpose of the researchMultiple myeloma and plasmacytomas belong to a group of disorders, namely plasma cell dyscrasias and are identified by the presence of a monoclonal protein (M-protein). MALDI-TOF-mass spectrometry (MS) has demonstrated superior analytical sensitivity for the detection of M-protein and is now used for screening of M-protein at some centres. We present the results of an alternative methodology for M-protein analysis by MALDI-TOF MS. MethodsSerum samples from patients with newly diagnosed multiple myeloma or plasmacytoma with positive M-protein detected by serum protein electrophoresis, immunofixation electrophoresis and serum free light chain analysis, underwent direct reagent-based extraction process using Acetonitrile (ACN) precipitation. Serum{kappa} and{lambda} light chains were validated using immunoenrichment by anti-{kappa} and anti-{lambda} biotin-labelled antibodies immobilised on streptavidin magnetic beads. MALDI-TOF MS measurements were obtained for intact proteins using alpha-cyano-4-hydroxycinnamic acid as matrix. The images obtained were overlaid on apparently healthy donor serum samples to confirm the presence of M-protein. Principle resultsCharacteristic M-protein peaks were observed in the ACN precipitates of serum in the predicted mass ranges for{kappa} and{lambda} . The{kappa} and{lambda} peaks were confirmed by immunoenrichment analysis. Twenty-seven patient samples with either newly diagnosed multiple myeloma or plasmacytoma with monoclonal gammopathy detected by the standard methods were chosen for Acetonitrile precipitation and analysed by MALDI-TOF MS. All 27 patient samples demonstrated a peak suggestive of M-protein with mass/charge (m/z) falling within the{kappa} and{lambda} range. The concordance rate with serum immunofixation electrophoresis and free light chain analysis was above 90%. Major conclusionsWe report the results of a low-cost reagent-based extraction process using Acetonitrile precipitation to enrich for{kappa} and{lambda} light chains, which can be used for the screening and qualitative analysis of M-protein.

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Mehra, N., Gopisetty, G., Subramani, J., Rajamanickam, A., Sundersingh, S., Karunakaran, P., Perumal Kalaiyarasi, J., Kannan, K., Radhakrishnan, V., Tenali Gnana, S., Thangarajan, R.. 2020-03-14. Detection of M-Protein in Acetonitrile Precipitates of Serum using MALDI-TOF Mass Spectrometry. https://doi.org/10.1101/2020.03.13.990192

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