Abstract: A method of separating free target analyte from protein-bound target analyte is described. Such can include obtaining an aqueous sample containing a target analyte in a free form (free target analyte) and the target analyte in a protein-bound form (protein-bound target analyte), passing the aqueous sample through a size exclusion chromatography matrix with a molecular weight cut off sufficient to allow the free target analyte to permeate into pores of the size exclusion chromatography matrix and exclude the protein-bound target analyte, whereupon the free target analyte adheres to and is immobilized by the size exclusion chromatography matrix and the protein-bound target analyte does not adhere to by the size exclusion chromatography matrix, separating the free target analyte from the protein-bound analyte by removing the protein-bound target analyte from the size exclusion chromatography matrix, and eluting the free target analyte from the size exclusion chromatography matrix with an organic solvent.

Abstract: Disclosed herein are methods of determining a concentration of IGF-1 and/or IGF-2 with improved sensitivity using high performance liquid chromatography (HPLC) and mass spectrometry (MS). The methods can achieve improved sensitivity by using a supercharging reagent as part of a mobile phase of the HPLC, which can increase the amount of specific charge states of IGF-1 and/or IGF-2 ions detected by MS. An example method includes subjecting a sample to HPLC, wherein the HPLC comprises a mobile phase including a supercharging reagent and an organic acid; ionizing the sample to produce one or more ions detectable by mass spectrometry; determining amounts of IGF-1 ions in an 8+ charge state by mass spectrometry; and relating the amount of the determined IGF-1 ions in the 8+ charge state to the concentration of IGF-1 in the sample.

Abstract: Methods for identifying and determining an abundance of an alpha fetoprotein (AFP) glycoform in a biological sample includes obtaining a biological sample, separating AFP from the biological sample as an enriched AFP sample, separating intact AFP from the enriched AFP sample, subjecting the intact AFP to high resolution mass spectrometry to generate a mass spectrum, identifying at least one AFP glycoform from the mass spectrum, and determining an abundance of the at least one AFP glycoform in the biological from the mass spectrum.

Abstract: A method of separating free target analyte from protein-bound target analyte is described. Such can include obtaining an aqueous sample containing a target analyte in a free form (free target analyte) and the target analyte in a protein-bound form (protein-bound target analyte), passing the aqueous sample through a size exclusion chromatography matrix with a molecular weight cut off sufficient to allow the free target analyte to permeate into pores of the size exclusion chromatography matrix and exclude the protein-bound target analyte, whereupon the free target analyte adheres to and is immobilized by the size exclusion chromatography matrix and the protein-bound target analyte does not adhere to by the size exclusion chromatography matrix, separating the free target analyte from the protein-bound analyte by removing the protein-bound target analyte from the size exclusion chromatography matrix, and eluting the free target analyte from the size exclusion chromatography matrix with an organic solvent.

Abstract: A method of separating free target analyte from protein-bound target analyte is described. Such can include obtaining an aqueous sample containing a target analyte in a free form (free target analyte) and the target analyte in a protein-bound form (protein-bound target analyte), passing the aqueous sample through a size exclusion chromatography matrix with a molecular weight cut off sufficient to allow the free target analyte to permeate into pores of the size exclusion chromatography matrix and exclude the protein-bound target analyte, whereupon the free target analyte adheres to and is immobilized by the size exclusion chromatography matrix and the protein-bound target analyte does not adhere to by the size exclusion chromatography matrix, separating the free target analyte from the protein-bound analyte by removing the protein-bound target analyte from the size exclusion chromatography matrix, and eluting the free target analyte from the size exclusion chromatography matrix with an organic solvent.

Abstract: The present invention provides methods, compositions, and kits associated with analyzing, enriching, and/or isolating a biomarker or analyte in a biological sample. In one aspect, for example, a method for determining a concentration of a biomarker in a biological sample can include binding any unbound biomarker with an antibody specific for the biomarker to form antibody-bound biomarker, enriching the antibody-bound biomarker and any endogenous autoantibody-bound biomarker to form an enriched fraction, identifying the biomarker in the enriched fraction, and determining the concentration of the biomarker in the biological sample. In one aspect, the concentration of the biomarker is derived from initially unbound biomarker and autoantibody-bound biomarker in the biological sample.

Abstract: Methods for measuring and analyzing parathyroid hormone-related peptide (PTHrP) using LC-MS/MS, including applications of the methods thereof, are disclosed and discussed. Such methods can include, along with the use of an isotope-labeled internal standard, purifying PTHrP from a biological sample, proteolytically digesting the PTHrP, and measuring specific digestion products using the using LC-MS/MS.

Abstract: A method of separating free target analyte from protein-bound target analyte is described. Such can include obtaining an aqueous sample containing a target analyte in a free form (free target analyte) and the target analyte in a protein-bound form (protein-bound target analyte), passing the aqueous sample through a size exclusion chromatography matrix with a molecular weight cut off sufficient to allow the free target analyte to permeate into pores of the size exclusion chromatography matrix and exclude the protein-bound target analyte, whereupon the free target analyte adheres to and is immobilized by the size exclusion chromatography matrix and the protein-bound target analyte does not adhere to by the size exclusion chromatography matrix, separating the free target analyte from the protein-bound analyte by removing the protein-bound target analyte from the size exclusion chromatography matrix, and eluting the free target analyte from the size exclusion chromatography matrix with an organic solvent.

Abstract: Methods for measuring and analyzing parathyroid hormone-related peptide (PTHrP) using LC-MS/MS, including applications of the methods thereof, are disclosed and discussed. Such methods can include, along with the use of an isotope-labeled internal standard, purifying PTHrP from a biological sample, proteolytically digesting the PTHrP, and measuring specific digestion products using the using LC-MS/MS.

Abstract: The present disclosure provides methods and composition involving increased sensitivity of compounds using mass spectrometry. In one embodiment, a method of increasing the sensitivity for detection of a carbonyl group-containing compound by mass spectrometry can comprise derivatizing the carbonyl group-containing compound with an O-substituted hydroxylamine thereby producing an oxime, resulting in enhanced sensitivity of detection by mass spectrometry, as compared to the underivatized carbonyl group-containing compound. Additionally, a method for assaying a carbonyl group-containing compound can comprise reacting the carbonyl group-containing compound in a sample with an O-substituted hydroxylamine to produce an oxime and performing analysis with mass spectrometric detection of the oxime by a mass spectrometry instrument.

Abstract: Methods for measuring and analyzing parathyroid hormone-related peptide (PTHrP) using LC-MS/MS, including applications of the methods thereof, are disclosed and discussed. Such methods can include, along with the use of an isotope-labeled internal standard, purifying PTHrP from a biological sample, proteolytically digesting the PTHrP, and measuring specific digestion products using the using LC-MS/MS.

Abstract: The present invention provides methods, compositions, and kits associated with analyzing, enriching, and/or isolating a biomarker or analyte in a biological sample. In one aspect, for example, a method for determining a concentration of a biomarker in a biological sample can include binding any unbound biomarker with an antibody specific for the biomarker to form antibody-bound biomarker, enriching the antibody-bound biomarker and any endogenous autoantibody-bound biomarker to form an enriched fraction, identifying the biomarker in the enriched fraction, and determining the concentration of the biomarker in the biological sample. In one aspect, the concentration of the biomarker is derived from initially unbound biomarker and autoantibody-bound biomarker in the biological sample.

Abstract: The present invention provides methods, compositions, and kits associated with analyzing, enriching, and/or isolating a biomarker or analyte in a biological sample. In one aspect, for example, a method for determining a concentration of a biomarker in a biological sample can include binding any unbound biomarker with an antibody specific for the biomarker to form antibody-bound biomarker, enriching the antibody-bound biomarker and any endogenous autoantibody-bound biomarker to form an enriched fraction, identifying the biomarker in the enriched fraction, and determining the concentration of the biomarker in the biological sample. In one aspect, the concentration of the biomarker is derived from initially unbound biomarker and autoantibody-bound biomarker in the biological sample.

Abstract: Concentrations of endogenous steroids in ovarian follicular fluid are used to develop steroid profiles which provide means for the diagnosis and prognosis of endocrine-related conditions and for identifying and developing appropriate treatments for related conditions, including the identification and development of suitable protocols for in vitro fertilization (IVF), treatment and predictive strategies for successful IVF outcomes and selected uses of oocytes for IVF or embryonic stem cell procedures.

Abstract: The present disclosure provides methods and composition involving increased sensitivity of compounds using mass spectrometry. In one embodiment, a method of increasing the sensitivity for detection of a carbonyl group-containing compound by mass spectrometry can comprise derivatizing the carbonyl group-containing compound with an O-substituted hydroxylamine thereby producing an oxime, resulting in enhanced sensitivity of detection by mass spectrometry, as compared to the underivatized carbonyl group-containing compound. Additionally, a method for assaying a carbonyl group-containing compound can comprise reacting the carbonyl group-containing compound in a sample with an O-substituted hydroxylamine to produce an oxime and performing analysis with mass spectrometric detection of the oxime by a mass spectrometry instrument.

Abstract: Concentrations of endogenous steroids in ovarian follicular fluid are used to develop steroid profiles which provide means for the diagnosis and prognosis of endocrine-related conditions and for identifying and developing appropriate treatments for related conditions, including the identification and development of suitable protocols for in vitro fertilization (IVF), treatment and predictive strategies for successful IVF outcomes and selected uses of oocytes for IVF or embryonic stem cell procedures.

Abstract: The present invention provides methods for high throughput analysis of analytes in complex mixtures for unresolved chromatographic peaks including specific embodiment for summing intensities for each mass transition of interest over a selected chromatographic peak (50) to generate a signal corresponding to total intensity for each transition (55, 60). The intensities are deconvoluted into intensities of individual analytes (65, 70), based on branching ratios acquired from authentic standards, and a comparison to calibration curve is performed to obtain a quantitative concentration measurement of a particular analyte in a sample.