title: Quantitative measurements of GDF-8 using immunoaffinity LC-MS/MS.
abstract: Growth and differentiation factor 8 (GDF-8) is a negative regulator of skeletal muscle mass and targeted by inhibitors to treat diseases associated with muscle loss. In order to enable clinical and translational investigations of GDF-8 inhibitors, specific and sensitive measurements of GDF-8 are necessary.
  
  
  
  An immunoaffinity LC-MS/MS assay for quantification of GDF-8 in serum was developed, qualified and implemented. The workflow includes offline enrichment of GDF-8 using an anti-GDF-8 antibody, followed by isolation using magnetic beads, trypsin digestion, and quantification using 2D nanoflow LC-MS/MS.
  
  
  
  This assay was qualified in human serum with a lower LOQ of 1.0 ng/mL based on the intact protein. GDF-8 was quantified in serum from juvenile and adult humans as well as mouse, rat, and cynomolgus monkey. Additionally, the assay was utilized to demonstrate an increase of total GDF-8 in serum following administration of an anti-GDF-8 monoclonal antibody therapeutic in cynomolgus monkeys.
  
  
  
  A specific and sensitive method was developed for the measurement of GDF-8 in juvenile and adult serum samples as well as preclinical species. The confident quantification of GDF-8 now enables a greater understanding of any association between changes GDF-8 levels and muscle mass.
authors: Pfizer Worldwide Research & Development, Andover, MA, USA.
authors: Joe
authors: J
authors: Palandra
authors: Pfizer Worldwide Research & Development, Cambridge, MA, USA.
authors: Amira
authors: A
authors: Quazi
authors: Pfizer Worldwide Research & Development, Cambridge, MA, USA.
authors: Lori
authors: L
authors: Fitz
authors: Pfizer Worldwide Research & Development, Cambridge, MA, USA.
authors: Haojing
authors: H
authors: Rong
authors: Pfizer Worldwide Research & Development, Cambridge, MA, USA.
authors: Carl
authors: C
authors: Morris
authors: Pfizer Worldwide Research & Development, Andover, MA, USA.
authors: Hendrik
authors: H
authors: Neubert
mesh_terms: D000595:Amino Acid Sequence; D000818:Animals; D000911:Antibodies, Monoclonal / Q000737:chemistry*; D001681:Biological Assay / Q000592:standards*; D002846:Chromatography, Affinity / Q000379:methods; D002853:Chromatography, Liquid / Q000379:methods; D015870:Gene Expression; D006801:Humans; D018189:Immunomagnetic Separation / Q000379:methods; D008252:Macaca fascicularis; D051379:Mice; D055435:Myostatin / Q000097:blood* / Q000235:genetics; D010455:Peptides / Q000138:chemical synthesis; D051381:Rats; D053719:Tandem Mass Spectrometry / Q000379:methods
chemical_list: D000911:Antibodies, Monoclonal; D055435:Myostatin; D010455:Peptides
keywords: Biomarkers; GDF-8; LC-MS/MS; immunoaffinity; nanospray; protein quantitation