Abstract: Provided are methods of detecting the presence or amount of NPY 1-36 or NPY 3-36 in a sample using mass spectrometry. Importantly, by methods of the invention, both NPY 1-36 and NPY 3-36 can be quantified simultaneously, separately, and independently in a sample that contains both peptides. The methods provide enhanced specificity, and excellent sensitivity with limits of quantitation (LOQ) of about 0.1 ng/mL, and are accomplished in less time and with less sample preparation than required in other assays for NPY. In certain embodiments, because the methods of the invention are specific for both NPY 1-36 and NPY 3-36, the methods provide a major advantage over existing immunoassays. Therefore, the methods may be used to obtain reliable concentration data in several important patient populations, such as patients with hypertension, heart disease, or cancer.

Abstract: Provided are methods of detecting the presence or amount of a dihydroxyvitamin D metabolite in a sample using mass spectrometry. The methods generally comprise associating an amine with a dihydroxyvitamin D metabolite in a sample, ionizing the adduct, and detecting the amount of the ion to determine the presence or amount of the vitamin D metabolite in the sample.

Abstract: The invention provides methods for simultaneously detecting or simultaneously quantifying any combination of thyroxine (T4), triiodothyronine (T3), 3,3?-diiodo-L-thyronine (3,3?-T2), 3-iodothyronamine (T1AM), and, optionally, reverse T3 (rT3) in a sample obtained from a human. The method involves a simple, sensitive, accurate, and specific isotope dilution tandem mass spectrometry method for the simultaneous quantification of any combination of T4, T3, 3,3?-T2, T1AM, and, optionally, rT3 in a sample obtained from a human, e.g., in human plasma or serum samples. This assay is far more sensitive than previously described assays for thyronamines and allows quantitation of T1AM in human plasma or serum, including from healthy controls.

Abstract: The invention provides methods for simultaneously detecting or simultaneously quantifying any combination of thyroxine (T4), triiodothyronine (T3), 3,3?-diiodo-L-thyronine (3,3?-T2), 3-iodothyronamine (T1AM), and, optionally, reverse T3 (rT3) in a sample obtained from a human. The method involves a simple, sensitive, accurate, and specific isotope dilution tandem mass spectrometry method for the simultaneous quantification of any combination of T4, T3, 3,3?-T2, T1AM, and, optionally, rT3 in a sample obtained from a human, e.g., in human plasma or serum samples. This assay is far more sensitive than previously described assays for thyronamines and allows quantitation of T1AM in human plasma or serum, including from healthy controls.

Abstract: Provided are methods of detecting the presence or amount of NPY 1-36 or NPY 3-36 in a sample using mass spectrometry. Importantly, by methods of the invention, both NPY 1-36 and NPY 3-36 can be quantified simultaneously, separately, and independently in a sample that contains both peptides. The methods provide enhanced specificity, and excellent sensitivity with limits of quantitation (LOQ) of about 0.1 ng/mL, and are accomplished in less time and with less sample preparation than required in other assays for NPY. In certain embodiments, because the methods of the invention are specific for both NPY 1-36 and NPY 3-36, the methods provide a major advantage over existing immunoassays. Therefore, the methods may be used to obtain reliable concentration data in several important patient populations, such as patients with hypertension, heart disease, or cancer.

Abstract: Provided are methods of detecting the presence or amount of a dihydroxyvitamin D metabolite in a sample using mass spectrometry. The methods generally comprise associating an amine with a dihydroxyvitamin D metabolite in a sample, ionizing the adduct, and detecting the amount of the ion to determine the presence or amount of the vitamin D metabolite in the sample.

Abstract: Methods, systems and kits for the simultaneous or sequential analysis of one or more hormones by mass spectrometry are disclosed. The methods require minimal sample size and minimal preparation time. The methods comprise ionizing the hormones and analyzing the hormones by mass spectrometry. In addition, methods, systems and kits for the simultaneous or sequential analysis of free thyroxine (FT4) hormone and free-triiodothyronine (FT3) is disclosed comprising ionization of the FT4 and FT3 hormone in the negative mode using an electrospray source.

Abstract: Methods, systems and kits for the simultaneous or sequential analysis of a multitude of steroid hormones by mass spectrometry are disclosed. The methods require minimal sample size and minimal preparation time. The methods include ionizing the hormones and analyzing the hormones by mass spectrometry. In addition, methods, systems and kits for the simultaneous or sequential analysis of steroid hormones are disclosed including ionization of the steroid hormones by photoionization.

Abstract: Methods, systems and kits for the simultaneous or sequential analysis of one or more hormones by mass spectrometry are disclosed. The methods require minimal sample size and minimal preparation time. The methods comprise ionizing the hormones and analyzing the hormones by mass spectrometry. In addition, methods, systems and kits for the simultaneous or sequential analysis of thyroid hormones are disclosed comprising ionization of the thyroid hormones in the negative mode using an electrospray source.

Abstract: Methods, systems and kits for the simultaneous or sequential analysis of one or more hormones by mass spectrometry are disclosed. The methods require minimal sample size and minimal preparation time. The methods comprise ionizing the hormones and analyzing the hormones by mass spectrometry. In addition, methods, systems and kits for the simultaneous or sequential analysis of steroid hormones are disclosed comprising ionization of the steroid hormones by photoionization.

Abstract: We have identified and purified to homogeneity from lymphatic tissues a novel 8.4 kDa immunophilin that specifically and avidly binds the immunosuppressant drugs FK-506 (Kd=0.8 nM) and rapamycin (Kd=0.08 nM) and their pharmacologically active metabolites and derivatives, but does not bind cyclosporin A. The isolated 8.4 kDa protein was analyzed for partial amino acid sequence, molecular weight, binding constants, binding specificity, biochemical aspects, and utility as the protein binding reagent in binding assays for immunosuppressant drugs in fluid samples, including patient blood.

Abstract: We have identified and purified to homogeneity from lymphatic tissues a 8.4 kDa immunophilin that specifically and avidly binds the immunosuppressant drugs FK-506 (Kd=0.8 nM) and rapamycin (Kd=0.08 nM) and their pharmacologically active metabolites and derivatives, but does not bind cyclosporin A. The isolated 8.4 kDa protein appears to be identical to authentic human and bovine ubiquitins in all measured respects (partial amino acid sequence, molecular weight, binding constants, binding specificity, biochemical aspects, and utility as the protein binding reagent in binding assays for immunosuppressant drugs in fluid samples, including patient blood). The availability of commercial quantities of human recombinant ubiguitin removes a supply barrier to the use of immunophilin protein binding assays for the estimation of FK-506, rapamycin and pharmacologically active metabolites and derivatives in the clinical setting.

Abstract: Purified immunosuppressive drug binding protein (immunophilin) of molecular weight 34-37 kDa and pI of about 6.5 is described. The 34-37 kDa immunophilin specifically binds FK-506, rapamycin and CsA with high affinity. This novel immunophilin can be used as a reagent for capturing, detecting and quantififying immunosuppressive drugs and their biologically active metabolites, derivatives and analogues in tissue or fluid samples, and for the capturing potential immunosuppressive drugs from microbial extracts or culture media.

Abstract: Purified immunosuppressive drug binding proteins (immunophilins) of molecular mass 2.4-3.0 kDa, 4.5 kDa, 34-37 kDa, 50-54 kDa, 80-100 kDa, and greater than about 120 kDa are described. The 34-37 kDa immunophilin specifically binds FK-506 and rapamycin. The 50-54 kDa immunophilin specifically binds FK-506, rapamycin and cyclosporine A, but with binding site distinctions. The 50-54 kDa immunophilin is devoid of significant rotomase activity, but inhibits cAMP-activated protein kinase activity. The amino acid composition, and the sequences of a dodecameric amino acid C-terminus partial sequence and of two heptameric internal partial amino acid sequences, of the 50-54 kDa immunophilin are described; the deduced molecular weight is 52,171. Recombinant about 52 kDa immunophilin is also described.

Abstract: A new homogeneous cytosolic binding protein (FKBP-14.6 immunophilin) having a molecular weight of about 14.6 kDa (calculated from its amino acid composition), reversibly binds the immunosuppressive drugs FK-506, rapamycin or chemically related compounds, but not cyclosporine. The FKBP-14.6 protein has a pI of 6.5-7.5, and the (partial) N-terminal amino acid sequence: NH.sub.2 -Lys-Leu-Pro-Tyr-Glu-Leu-Lys-X-Asn-Val-Lys-Ala-Phe-X-X-Lys-Val- (where X is undefined), and partial internal amino acid sequences -Val-Leu-Asp-Thr-Ala-Tyr-Glu-Tyr-Gly-Ala-Glu-Ala-Leu-Glu-, -Glu-Phe-Thr-Pro-Val-Phe-Gln-Ala-X-Phe- and -Ser-Leu-Val-Pro-Leu-Val-Gly-X-Lys- (where X is undefined). This N-terminal amino acid sequence exhibits no homology to FKBP-12 or membrane-associated FKBP-13. THe FKBP-14.6 is isolated from the cytosol of mammalian tissues, preferably calf thymus, and can be used in diagnostic and purification procedures involving FK-506 and rapamycin-type immunosuppressant drugs.

Abstract: A new homogeneous cytosolic binding protein (FKBP-14.6 immunophilin) having a molecular weight of about 14.6 kDa (calculated from its amino acid composition), reversibly binds the immunosuppressive drugs FK-506, rapamycin or chemically related compounds, but not cyclosporine. The FKBP-14.6 protein has a pI of 6.5-7.5, and the (partial) N-terminal amino acid sequence: NH.sub.2 -Lys-Leu-Pro-Tyr-Glu-Leu-Lys-Xaa-Asn-Val-Lys-Ala-Phe-Xaa-Xaa-Lys-Val- (Seq. ID. No: 1) (where Xaa is undefined), and partial internal amino acid sequences -Val-Leu-Asp-Thr-Ala-Tyr-Glu-Tyr-Gly-Ala-Glu-Ala-Leu-Glu- (Seq. ID. No: 2), -Glu-Phe-Thr-Pro-Val-Phe-Gln-Ala-Xaa-Phe- (Seq. ID. No: 3) and -Ser-Leu-Val-Pro-Leu-Val-Gly-Xaa-Lys- (Seq. ID. No. 4) (where Xaa is undefined). This N-terminal amino acid sequence exhibits no homology to FKBP-12 or membrane-associated FKBP-13. The FKBP-14.

Abstract: A competitive protein binding assay for rapamycin and biologically-active metabolites, derivatives and analogues thereof in blood and other biological fluids is disclosed, wherein the binding reagent is a specific rapamycin binding protein either substantially purified from the soluble cytoplasm of target cells of rapamycin action, particularly normal or transformed lymphocytes, freshly collected or in established cell lines, or synthesized by recombinant DNA techniques. Solution phase and solid state assay systems are disclosed.

Abstract: Methods, systems and kits for the simultaneous or sequential analysis of one or more hormones by mass spectrometry are disclosed. The methods require minimal sample size and minimal preparation time. The methods include ionizing the hormones and analyzing the hormones by mass spectrometry. In addition, methods, systems and kits for the simultaneous or sequential analysis of thyroid hormones are disclosed including ionization of the thyroid hormones in the negative mode using an electrospray source.