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  • Beck, O., et al. (författare)
  • Potential of Mass Spectrometry in Developing Clinical Laboratory Biomarkers of Nonvolatiles in Exhaled Breath
  • 2016
  • Ingår i: Clinical Chemistry. - : Oxford University Press (OUP). - 0009-9147 .- 1530-8561. ; 62:1, s. 84-91
  • Tidskriftsartikel (refereegranskat)abstract
    • BACKGROUND: Exhaled breath contains nonvolatile substances that are part of aerosol particles of submicrometer size. These particles are formed and exhaled as a result of normal breathing and contain material from distal airways of the respiratory system. Exhaled breath can be used to monitor biomarkers of both endogenous and exogenous origin and constitutes an attractive specimen for medical investigations. CONTENT: This review summarizes the present status regarding potential biomarkers of nonvolatile compounds in exhaled breath. The field of exhaled breath condensate is briefly reviewed, together with more recent work on more selective collection procedures for exhaled particles. The relation of these particles to the surfactant in the terminal parts of the respiratory system is described. The literature on potential endogenous low molecular weight compounds as well as protein biomarkers is reviewed. The possibility to measure exposure to therapeutic and abused drugs is demonstrated. Finally, the potential future role and importance of mass spectrometry is discussed. SUMMARY: Nonvolatile compounds exit the lung as aerosol particles that can be sampled easily and selectively. The clinical applications of potential biomarkers in exhaled breath comprise diagnosis of disease, monitoring of disease progress, monitoring of drug therapy, and toxicological investigations. (C) 2015 American Association for Clinical Chemistry
  • Berg, Cecilia, 1969-, et al. (författare)
  • Direct solid-phase sequence analysis of the human p53 gene by use of multiplex polymerase chain reaction and alpha-thiotriphosphate nucleotides.
  • 1995
  • Ingår i: Clinical Chemistry. - : Oxford University Press (OUP). - 0009-9147 .- 1530-8561. ; 41:10, s. 1461-6
  • Tidskriftsartikel (refereegranskat)abstract
    • Among the candidate cancer-prognostic genes is the p53 tumor suppressor gene, which, when mutated, plays an important role in the development of many types of cancers. To facilitate robust large-scale DNA analysis of microdissected tumor biopsies, we describe a multiplex/nested PCR approach for a simultaneous outer amplification of exons 4-9 of the human p53 gene with parallel amplification of the HLA-DQB1 locus, involving a total of 14 primers. This approach reduces the required number of cells for analysis and avoids any variation in the amplifications of the individual p53 exons during the common outer amplification step. The HLA sequencing allows sample identification because the DQB1 locus is highly polymorphic and is thereby patient-specific. The p53 and HLA amplicons are analyzed by solid-phase sequencing in a semiautomated format. To improve the DNA sequence quality, we used 2'-deoxyribonucleoside 5'-O-1-thiotriphosphates in the sequencing reactions.
  • Berois, Nora, et al. (författare)
  • ppGalNAc-TI3 : A new molecular marker of bone marrow involvement in neuroblastoma
  • 2006
  • Ingår i: Clinical Chemistry. - : Oxford University Press (OUP). - 0009-9147 .- 1530-8561. ; 52:9, s. 1701-1712
  • Tidskriftsartikel (refereegranskat)abstract
    • Background: To identify new molecular markers of bone marrow dissemination in human neuroblastoma (NB), we studied the transcriptome profiles of malignant neuroblasts established from the human MYCN-amplified IGR-N-91 model. Methods: This experimental model includes human neuroblastoma cells derived from & subcutaneous stage 4 disease, myocardium (Myoc) and bone marrow (BM) metastatic cells. Results: Gene expression profiles obtained with Agilent oligo microarrays revealed a set of 107 differentially expressed genes in the metastatic neuroblasts. This set included up-regulated genes involved in chemoresistance, cell motility, neuronal structure/signaling, and the recently characterized GALNT13 gene encoding a glycosyltransferase that initiates mucin-type O-glycosylation. Because the glycosylation process is involved in the progression of primary tumor to metastatic disease, we investigated whether the most strongly upregulated gene, GALNT13, might be a marker of bone marrow involvement in stage 4 NB patients. Importantly, in the BM of healthy adults no GALNT13 transcript was detected with analysis by quantitative (n = 3) and nested reverse transcription-PCR (n = 4) assays. In contrast, GALNT13 transcripts were detected in 23/23 cytologically involved BM samples obtained at diagnosis of stage 4 NB patients and in 5/27 cytologically noninvolved BM samples obtained from patients with stage 1-4 and 4S and treated stage 4 NB. The quantitative measurements of tyrosine hydroxylase (TH), ganglioside D2 synthase, dopa decarboxylase, and GALNT13 transcript values were compared in the same NB patients, and the results showed that GALNT13 expression was most highly correlated to poor clinical outcome at diagnosis. Conclusion: We propose ppGalNAc-T13 as a new informative marker for the molecular diagnosis of BM involvement and the follow-up of minimal residual disease in NB patients. © 2006 American Association for Clinical Chemistry.
  • Bredberg, Anna, et al. (författare)
  • Exhaled Endogenous Particles Contain Lung Proteins.
  • 2012
  • Ingår i: Clinical Chemistry. - : Oxford University Press (OUP). - 1530-8561 .- 0009-9147. ; 58:2, s. 431-440
  • Tidskriftsartikel (refereegranskat)abstract
    • BACKGROUND:We recently developed a novel, noninvasive method for sampling nonvolatile material from the distal airways. The method is based on the collection of endogenous particles in exhaled air (PEx). The aim of this study was to characterize the protein composition of PEx and to verify that the origin of PEx is respiratory tract lining fluid (RTLF).METHOD:Healthy individuals exhaled into the sampling device, which collected PEx onto a silicon plate inside a 3-stage impactor. After their extraction from the plates, PEx proteins were separated by SDS-PAGE and then analyzed by LC-MS. Proteins were identified by searching the International Protein Index human database with the Mascot search engine.RESULTS:Analysis of the pooled samples identified 124 proteins. A comparison of the identified PEx proteins with published bronchoalveolar lavage (BAL) proteomic data showed a high degree of overlap, with 103 (83%) of the PEx proteins having previously been detected in BAL. The relative abundances of the proteins were estimated according to the Mascot exponentially modified protein abundance index protocol and were in agreement with the expected protein composition of RTLF. No amylase was detected, indicating the absence of saliva protein contamination with our sampling technique.CONCLUSIONS:Our data strongly support that PEx originate from RTLF and reflect the composition of undiluted RTLF.
  • Carlsson, Sigrid, 1982, et al. (författare)
  • Perspective on prostate cancer screening
  • 2019
  • Ingår i: Clinical Chemistry. - : Oxford University Press (OUP). - 0009-9147 .- 1530-8561. ; 65:1, s. 24-27
  • Tidskriftsartikel (refereegranskat)
  • Ceder, Gunnel, et al. (författare)
  • Concentration ratios of morphine to codeine in blood of impaired drivers as evidence of heroin use and not medication with codeine
  • 2001
  • Ingår i: Clinical Chemistry. - 0009-9147 .- 1530-8561. ; 47:11, s. 1980-1984
  • Tidskriftsartikel (refereegranskat)abstract
    • Background: Both the illicit drug heroin and the prescription drug codeine are metabolized to morphine, which tends to complicate interpretation of opiate-positive samples. We report here the concentrations of morphine and codeine, the morphine/codeine ratios, and 6-acetylmorphine (6-AM) in blood specimens from individuals arrested for driving under the influence of drugs (DUID) in Sweden. The results were compared with positive findings of 6-AM in urine as evidence of heroin intake. Methods: In 339 DUID suspects, both blood and urine specimens were available for toxicologic analysis. In another 882 cases, only blood was available. All specimens were initially analyzed by immunoassay, and the positive results were verified by isotope-dilution gas chromatography-mass spectrometry. In routine casework, the limits of quantification (LOQs) for unconjugated opiates were 5 ng/g for blood and 20 ╡g/L for urine. Results: The median concentration of morphine in blood was 30 ng/g with 2.5 and 97.5 percentiles of 5 and 230 ng/g, respectively (n = 979). This compares with a median codeine concentration of 20 ng/g and 2.5 and 97.5 percentiles of 5 and 592 ng/g, respectively (n = 784). The specific metabolite of heroin, 6-AM, was identified in only 16 of 675 blood specimens (2.3%). This compares with positive findings of 6-AM in 212 of 339 urine samples (62%) from the same population of DUID suspects. When 6-AM was identified in urine, the morphine/codeine ratio in blood was always greater than unity (median, 6.0, range, 1-66). In 18 instances, 6-AM was present in urine, although morphine and codeine were below the LOQ in blood. The morphine/codeine ratio in blood was greater than unity in 85% of DUID cases when urine was not available (n = 506), and the median morphine and codeine concentrations were 70 ng/g and 10 ng/g, respectively. When morphine/codeine ratios in blood were less than unity (n = 76), the median morphine and codeine concentrations were 10 ng/g and 180 ng/g, respectively. Conclusions: Only 2.3% of opiate-positive DUID suspects were verified as heroin users on the basis of positive findings of 6-AM in blood. A much higher proportion (62%) were verified heroin users from 6-AM identified in urine. When urine was not available for analysis, finding a morphine/codeine concentration ratio in blood above unity suggests heroin use and not medication with codeine. This biomarker indicated that 85% of opiate-positive DUID blood samples were from heroin users. ⌐ 2001 American Association for Clinical Chemistry.
  • Chalbot, Sonia, et al. (författare)
  • Cerebrospinal fluid secretory Ca2+-dependent phospholipase A2 activity is increased in Alzheimer disease.
  • 2009
  • Ingår i: Clinical chemistry. - : Oxford University Press (OUP). - 1530-8561 .- 0009-9147. ; 55:12, s. 2171-9
  • Tidskriftsartikel (refereegranskat)abstract
    • BACKGROUND: The phospholipase A(2) (PLA2) family comprises multiple isoenzymes that vary in their physicochemical properties, cellular localizations, calcium sensitivities, and substrate specificities. Despite these differences, PLA2s share the ability to catalyze the synthesis of the precursors of the proinflammatory mediators. To investigate the potential of PLA2 as a biomarker in screening neuroinflammatory disorders in both clinical and research settings, we developed a PLA2 assay and determined the predominant types of PLA2 activity in cerebrospinal fluid (CSF). METHODS: We used liposomes composed of a fluorescent probe (bis-Bodipy FL C11-PC [1,2-bis-(4,4- difluoro-5,7-dimethyl-4-bora-3a,4a-diaza-s-indacene-3-undecanoyl)-sn-glycero-3-phosphocholine]) and 1,2-dioleoyl-l-alpha-phosphatidylcholine as a substrate to measure CSF PLA2 activity in a 96-well microtiter plate format. We established the type of CSF PLA2 activity using type-specific inhibitors of PLA2. RESULTS: Using 5 microL CSF per assay, our PLA2 activity assay was reproducible with CVs <15% in 2 CSF samples and for recombinant secretory Ca(2+)-dependent PLA2 (sPLA2) in concentrations ranging from 0.25 to 1 micromol/L. This PLA2 assay allowed identification of sPLA2 activity in lumbar CSF from healthy individuals 20-77 years old that did not depend on either sex or age. Additionally, CSF sPLA2 activity was found to be increased (P = 0.0008) in patients with Alzheimer disease. CONCLUSIONS: Adult human CSF has sPLA2 activity that can be measured reliably with the assay described. This enzyme activity in the CSF is independent of both sex and age and might serve as a valuable biomarker of neuroinflammation, as we demonstrated in Alzheimer disease.
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