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Biotech / Medical : Ciphergen Biosystems(CIPH): -- Ignore unavailable to you. Want to Upgrade?

To: tuck who wrote (492)	4/9/2007 1:04:31 PM
From: tuck	Respond to of 510

>>Effects of protein stabilizers and storage time on the mass spectrometry dynamics of human serum proteins Presentation Start/End Time: Monday, Apr 16, 2007, 1:00 PM - 5:00 PM Location: Exhibit Hall, Los Angeles Convention Center Poster Section: 25 Poster Board Number: 28 Author Block: Amit S. Dhamoon, Rick Saul, Chenwei Liu, Elise C. Kohn, Gordon Whiteley. National Institutes of Health, Bethesda, MD, SAIC-Frederick, Clinical Proteomics Reference Laboratory, Gaithersburg, MD Optimization of pre-analytical variables in serum proteomics research is necessary to further develop this technology for biomarker discovery. We hypothesized that the presence of protein stabilizers (PS) in serum collection tubes at different storage times would affect the quality of mass-spectrometry (MS) data. We analyzed the effect of these variables on putative biomarkers for ovarian cancer (transthyretin variants and ApoA1) and on possible phosphorylation events measured by a novel bioinformatics method. Serum from 20 apparently healthy volunteers was collected with and without PS (BD P200 tube, Becton, Dickinson & Co; PS components proprietary). Serum was then frozen (-76 oC) immediately after preparation (t=0), or after 24, 48, or 72 hours of storage at 4 oC. Serum was fractionated using strong anion exchanger Ciphergen Q10 chips, and was studied using the Ciphergen SELDI platform. Separate aliquots of serum were fractionated using PerkinElmer Proexpression kits, which concentrate albumin-bound peptides, and studied using a PerkinElmer PrOTOF MS. Paired data were compared at each time point and repeated measure analysis was utilized to determine data trends over time. Total ionic current (TIC), as a surrogate of total protein load, was lower in serum tubes with PS. TIC measured on the Ciphergen MS was stable over time, while TIC on the PrOTOF MS decreased (p<0.05) over time in both tube types. Intensities of ApoA1 (m/z=28000), and unmodified (m/z=13736) and cysteinylated (m/z=13858) transthyretin peaks were statistically lower in tubes with PS at each time point (p<0.02, p<0.0002, p<0.015, respectively). Conversely, intensity of glutathionylated transthyretin (m/z=14050) was consistently higher (p<0.0003) in tubes with PS. With increasing storage time, intensity of unmodified transthyretin decreased (p<0.0001) in both tube types while intensities of ApoA1 and cysteinylated transthyretin did not show a significant change. Truncated transthyretin (m/z=12795) decreased (p=0.0003) without PS and glutationylated transthyretin increased (p=0.0025) with PS over greater storage time. PrOTOF datastreams were evaluated and 6 un- and putatively monophosphorylated peptides, or peak pairs, were identified by MS mass shifts of approximately 80 Da. Statistically significant differences in these peptide phosphorylation pairs between the two tube types were identified in 5 of 6 peak pairs. The presence of PS significantly alters the serum proteome over time as examined by SELDI and MALDI platforms. It is unclear how the presence of PS in serum collection tubes will impact the search for clinically useful biomarkers of disease. Further investigation is required to determine whether PS should be universally implemented in serum proteomics research.<< Both of these underscore the difficulty SELDI still faces as a diagnostic tool. Various things have been tried with mixed results. Here protein stabilizers. There the filtering out of low abundance proteins. The above study is particularly interesting because it discusses a couple of the biomarkers CIPH is using in its soon to launched (yeah, right) ovarian cancer test. And as one can see, protein stabilizers mess up the biomarkers, and not uniformly. This might explain CIPH's 25% slide in the last couple of weeks. Or it might be the patent re-examination mentioned in the recent 10-K, which isn't going well. CIPH could lose $2 million from Bio-Rad, and it could lose exclusivity in detecting single biomarkers with SELDI. My guess is Qiagen, who bought LumiCyte, is behind this. Cheers, Tuck

To: tuck who wrote (492)	5/7/2007 10:31:48 AM
From: tuck	Respond to of 510

[Influence of blood sampling on protein profiling and pattern analysis using MALDI-MS.] >>BJU Int. 2007 Mar;99(3):658-62. Influence of blood sampling on protein profiling and pattern analysis using matrix-assisted laser desorption/ionisation mass spectrometry. Pelzer AE, Feuerstein I, Fuchsberger C, Ongarello S, Bektic J, Schwentner C, Klocker H, Bartsch G, Bonn GK. Department of Urology, Innsbruck Medical University, Anichstrasse 35, 6020-Innsbruck, Austria. alexandre.pelzer@uibk.ac.at OBJECTIVE: To describe the influence of blood sampling/sampling tubes on mass spectrometric and clustering results, and on clinical blood variables, in blood samples collected from healthy volunteers and patients with prostate cancer. PATIENTS, SUBJECTS AND METHODS: Two venous blood samples were taken from 12 healthy volunteers and 12 patients with localized prostate cancer. Two blood samples were taken from each participant using two different venepuncture systems (group A and group B). The Kolmogorov-Smirnov test was used to identify the peaks distinguishing the different groups. In a 10-fold cross-validation study, decision trees for identifying discriminatory peaks that separate the benign from the malignant were constructed. RESULTS: The decision tree separated samples measured by matrix-assisted laser desorption/ionization mass spectrometry (MALDI MS) from healthy volunteers from those of patients with prostate cancer, with a sensitivity of 93.6% and a specificity of 91.6%. Of special interest is that one peak at 6941 m/z was produced during blood sample preparation and had a very powerful influence on the results of the classification. CONCLUSION: The results clearly showed that blood-sampling systems have a great influence on the recorded MALDI MS traces, and thus can markedly influence and confound the results of the MS analysis, whereas clinical variables might remain unchanged. MS profiling is a promising method of marker discovery, but as it could be shown well-designed studies are critical to allow proper interpretation for the identification of key variables as well as for the clinical use.<< Well, Quest? Cheers, Tuck