Formalin-fixed isotope-labeled reference standards and methods for fabrication and use thereof

Abstract

One or more cells are labeled with minor stable isotopes, characterized, and preserved for subsequent use as a bio-specimen reference standard. The one or more cells are grown in culture media supplied with minor stable isotopes in concentrations substantially different from normally occurring concentrations, thereby supplanting major stable isotopes that would normally be incorporated into the proteins of the cells. The proteins of the cells are thus labeled by the minor stable isotopes and can be used in proteomic characterization of the cells. The cells are preserved by fixation as a reference standard. Cells of the reference standard are mixed with the sample and subject to mass spectrometry evaluation, whereby the labeled proteins of the reference standard can be used in determining the proteome of the sample.

1 . A method comprising:
 (a) for each of one or more cell lines, characterizing a respective proteome thereof using mass spectrometry;   (b) before or after (a), preserving the one or more cell lines by fixation so as to form one or more bio-specimen reference standards, each bio-specimen reference standard having proteins labeled with a minor stable isotope; and   (c) after (b), using the one or more bio-specimen reference standards to characterize a proteome of a sample.   
     
     
         2 . The method of  claim 1 , wherein:
 in (a), the one or more cell lines is a plurality of cell lines;   in (b), each cell line is preserved by fixation so as to form respective bio-specimen reference standards; and   (c) comprises selecting one of the bio-specimen references standards to analyze the sample, the selected bio-specimen reference standard being used to characterize the proteome of the sample.   
     
     
         3 . The method of  claim 2 , wherein the selecting is based at least in part on the respective characterized proteome. 
     
     
         4 . The method of  claim 1 , wherein (c) comprises assigning to the sample a protein composition based on a morphological feature of the sample that is shared with one or more bio-specimen reference standards. 
     
     
         5 . The method of  claim 1 , further comprising:
 analyzing the morphology of the one or more bio-specimen reference standards using at least one of microscopy, immunohistochemistry (IHC), immunofluorescence (IF), hematoxylin and eosin (H&E) staining; and   based on said analyzing and the characterized proteomes, selecting morphological features of the one or more bio-specimen reference standards that corresponds to protein compositions thereof.   
     
     
         6 . The method of  claim 5 , wherein (c) comprises assigning to the sample a protein composition based on a morphological feature of the sample that corresponds to one of the selected morphological features of the one or more bio-specimen reference standards. 
     
     
         7 . The method of  claim 1 , wherein:
 the one or more bio-specimen reference standards is a plurality of bio-specimen reference standards, and   (b) comprises subjecting each of the bio-specimen reference standards to a different fixation process so as to have varying levels or degrees of fixation.   
     
     
         8 . The method of  claim 1 , wherein each bio-specimen reference standard is formed of the respective cell line comprised of primary cells.