Analytical techniques for cell fractions

Anderson, Norman G.; Willis, D.D.; Holladay, D.W.; Caton, J.E.; Holleman, J.W.; Eveleigh, J. W.; Attrill, J.E.; Ball, Frances L.; Anderson, N. Leigh

doi:10.1016/0003-2697(75)90734-4

Cited by 23 publications

(8 citation statements)

References 2 publications

Supporting

Mentioning

Contrasting

Order By: Relevance

“…Stable isotope standard (SIS) monitor peptides were synthesized by Cell Signaling Technology Inc., (Beverley, MA) and each contained a single fully 13 C-substituted amino acid near the C-terminus (Table 3). The peptides were designated SIS 2-5 and corresponded to the natural peptides Nat 2-5.…”

Section: Labeled Peptidesmentioning

confidence: 99%

“…The peptides were designated SIS 2-5 and corresponded to the natural peptides Nat 2-5. Peptide SIS 3 (labeled on Phe) was 10 atomic mass units (AMU) heavier than the natural peptide (Nat 3), and the other SIS:Nat pairs differed by 6 AMU due to incorporation of a single 13 C-substituted Pro or Val. Peptides were quantitated by acid hydrolysis and amino acid analysis (AAA Services Inc., Boring, OR) for use as quantitative standards.…”

Section: Labeled Peptidesmentioning

confidence: 99%

“…Such an enrichment method can be applied routinely if the antibody adsorbent is capable of being recycled, a property which has been well-established, at least for polyclonal antibodies and protein ligands. 8,13,14 We have investigated an approach (summarized in Figure 1) that we refer to by the acronym SISCAPA (Stable Isotope Standards with Capture by Anti-Peptide Antibodies). SISCAPA combines the four elements previously described: (1) digestion of a protein sample to peptides; (2) addition of internal standard peptides labeled with stable isotopes; (3) enrichment of low abundance peptides by capture with immobilized antibodies; and (4) quantitation by MS.…”

Section: Introductionmentioning

confidence: 99%

See 2 more Smart Citations

Mass Spectrometric Quantitation of Peptides and Proteins Using Stable Isotope Standards and Capture by Anti-Peptide Antibodies (SISCAPA)

et al. 2004

View full text Add to dashboard Cite

A method (denoted SISCAPA) for quantitation of peptides in complex digests is described. In the method, anti-peptide antibodies immobilized on 100 nanoliter nanoaffinity columns are used to enrich specific peptides along with spiked stable-isotope-labeled internal standards of the same sequence. Upon elution from the anti-peptide antibody supports, electrospray mass spectrometry is used to quantitate the peptides (natural and labeled). In a series of pilot experiments, tryptic test peptides were chosen for four proteins of human plasma (hemopexin, alpha1 antichymotrypsin, interleukin-6, and tumor necrosis factor-alpha) from a pool of 10,203 in silico tryptic peptide candidates representing 237 known plasma components. Rabbit polyclonal antibodies raised against the chosen peptide sequences were affinity purified and covalently immobilized on POROS supports. Binding and elution from these supports was shown to provide an average 120-fold enrichment of the antigen peptide relative to others, as measured by selected ion monitoring (SIM) or selected reaction monitoring (SRM) electrospray mass spectrometry. The columns could be recycled with little loss in binding capacity, and generated peptide ion current measurements with cycle-to-cycle coefficients of variation near 5%. Anti-peptide antibody enrichment will contribute to increased sensitivity of MS-based assays, particularly for lower abundance proteins in plasma, and may ultimately allow substitution of a rapid bind/elute process for the time-consuming reverse phase separation now used as a prelude to online MS peptide assays. The method appears suitable for rapid generation of assays for defined proteins, and should find application in the validation of diagnostic protein panels in large sample sets.

show abstract

Section: Labeled Peptidesmentioning

confidence: 99%

Section: Labeled Peptidesmentioning

confidence: 99%

Section: Introductionmentioning

confidence: 99%

See 1 more Smart Citation

Mass Spectrometric Quantitation of Peptides and Proteins Using Stable Isotope Standards and Capture by Anti-Peptide Antibodies (SISCAPA)

et al. 2004

View full text Add to dashboard Cite

show abstract

“…The conventional proteomics technology platform most commonly used is based on 2-DE MS, which has been more successful in profiling proteins and their disease- or treatment- related quantitative changes in tissue homogenates than in plasma sample . Anderson have reviewed the past decades of plasma proteome research works based on 2-DE: ,− the highest quantity of identified plasma proteins is only 319. Despite inherent disadvantages of 2-DE, the reason of these relatively poor results maybe lies in the following aspects: the relatively serious band broadening due to very high abundance of a few proteins in plasma, and the crowding of separated protein spots in the molecular mass range between 45 and 80 KD and in the isoelectric point range between 4.5 and 6 …”

Section: Introductionmentioning

confidence: 99%

Human Plasma Proteome Analysis by Multidimensional Chromatography Prefractionation and Linear Ion Trap Mass Spectrometry Identification

Jin

Dai

et al. 2005

J. Proteome Res.

View full text Add to dashboard Cite

A resurgence of interest in the human plasma proteome has occurred in recent years because it holds great promise of revolution in disease diagnosis and therapeutic monitoring. As one of the most powerful separation techniques, multidimensional liquid chromatography has attracted extensive attention, but most published works have focused on the fractionation of tryptic peptides. In this study, proteins from human plasma were prefractionated by online sequential strong cation exchange chromatography and reversed-phase chromatography. The resulting 30 samples were individually digested by trypsin, and analyzed by capillary reversed-phase liquid chromatography coupled with linear ion trap mass spectrometry. After meeting stringent criteria, a total of 1292 distinct proteins were successfully identified in our work, among which, some proteins known to be present in serum in <10 ng/mL were detected. Compared with other works in published literatures, this analysis offered a more full-scale list of the plasma proteome. Considering our strategy allows high throughput of protein identification in serum, the prefractionation of proteins before MS analysis is a simple and effective method to facilitate human plasma proteome research.

show abstract

“…The strategy was to develop several different technologies in parallel. These included some of the first high pressure liquid chromatographic systems to resolve low molecular weight compounds [6] including sugars [7] and the constituents of nucleic acids [8], high resolution centrifuges to resolve cell components [9,10], and methods to separate proteins including rapid recycling immuno-affinity chromatographic systems to simplify complex protein mixtures [11,12]. Since it was realized that automatic fractionation methods would yield large numbers of samples to be analyzed for enzyme activities, the first computerized enzyme analyzer (the centrifugal fast analyzer) was invented, developed, and came into general use in clinical chemistry laboratories worldwide [13].…”

Section: History Of Molecular Anatomymentioning

confidence: 99%

Back to the future: The human protein index (HPI) and the agenda for post-proteomic biology

2001

View full text Add to dashboard Cite

The effort to produce an index of all human proteins (the human protein index, or HPI) began twenty years ago, before the initiation of the human genome program. Because DNA sequencing technology is inherently simpler and more scalable than protein analytical technology, and because the finiteness of genomes invited a spirit of rapid conquest, the notion of genome sequencing has displaced that of protein databases in the minds of most molecular biologists for the last decade. However, now that the human genome sequence is nearing completion, a major realignment is under way that brings proteins back to the center of biological thinking. Using an influx of new and improved protein technologies--from mass spectrometry to re-engineered two-dimensional (2-D) gel systems, the original objectives of the HPI have been expanded and the time frame for its execution radically shortened. Several additional large scale technology efforts flowing from the HPI are also described.

show abstract

Analytical techniques for cell fractions

Cited by 23 publications

References 2 publications

Mass Spectrometric Quantitation of Peptides and Proteins Using Stable Isotope Standards and Capture by Anti-Peptide Antibodies (SISCAPA)

Mass Spectrometric Quantitation of Peptides and Proteins Using Stable Isotope Standards and Capture by Anti-Peptide Antibodies (SISCAPA)

Human Plasma Proteome Analysis by Multidimensional Chromatography Prefractionation and Linear Ion Trap Mass Spectrometry Identification

Back to the future: The human protein index (HPI) and the agenda for post-proteomic biology

Contact Info

Product

Resources

About