New method to assess dilution of secretions for immunological and microbiological assays

Virolainen, Anni; Mäkelä, Mika; Esko, E.; Jero, Jussi; Alfthan, Georg; Sundvall, Jouko; Leinonen, Maija

doi:10.1128/jcm.31.5.1382-1384.1993

Cited by 11 publications

(4 citation statements)

References 11 publications

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“…Third, the Li cation is not capable of interaction with immunoglobulins or with immune complexes or of effective chemical binding with plasma proteins (4). Finally, even at concentrations of 20 meq/liter, Li interferes with neither EIA nor the survival of bacteria and viruses (6). Furthermore, we verified that the presence of LiCl at the final concentration of 10 mM did not affect the results of anti-HIV antibody detection in CVS with three EIAs based on different principles.…”

Section: Discussionsupporting

confidence: 51%

Dilution assessment of cervicovaginal secretions obtained by vaginal washing for immunological assays

Bélec

Meillet

Levy

et al. 1995

Clin Diagn Lab Immunol

103

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Local immunological defense mechanisms in the cervicovaginal mucosa currently remain incompletely defined, especially from a quantitative point of view. Addition of an inert substance, lithium chloride (LiCl), into the washing buffer used to carry out the vaginal washing for collecting cervicovaginal secretions and measurement of its concentration with a flame absorption spectrophotometer, before and after the specimen is sampled, permits the quantification of the volume of cervicovaginal secretions collected and the approximation of the dilution factor of a soluble component introduced by the washing. Lithium, at a concentration of 10 mM, gives the best precision of measurement and has no effect on the results of the immunoassays. In a population of 27 nonpregnant women (age range, 18 to 45 years), the volume of cervicovaginal secretions collected by vaginal washing with 3 ml of LiCl-phosphate-buffered saline was 12% ؎ 3.2% (mean ؎ standard deviation) of the total volume and showed large interindividual variations (range, 5.6 to 18.8%); the mean dilution factor of a soluble component from the vaginal secretions was 9.9% ؎ 2.8% (range, 6.3 to 18.8%). According to the date of the menstrual cycle, the mean volume of collected cervicovaginal secretions was significantly increased in the luteal phase in comparison with the follicular phase; conversely, the mean dilution factor of a soluble component was more important in the follicular than in the luteal phase. These features strengthen the need to quantify accurately the dilution factor introduced by vaginal washing when studying cervicovaginal immunity.

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Section: Discussionsupporting

confidence: 51%

Dilution assessment of cervicovaginal secretions obtained by vaginal washing for immunological assays

Bélec

Meillet

Levy

et al. 1995

Clin Diagn Lab Immunol

103

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“…Dilution effects associated with nasal lavage have also been addressed as a problem . This problem may be solved by normalizing against an endogenous dilution marker, e.g., albumin, total protein abundance, or urea. , Also, several exogenous dilution markers, e.g., inulin or lithium chloride, which are added to the washing fluid before sample collection, have been used . In this study, unknown dilution was corrected for by normalizing samples against total protein abundances.…”

Section: Discussionmentioning

confidence: 99%

Screening Method Using Selected Reaction Monitoring for Targeted Proteomics Studies of Nasal Lavage Fluid

et al. 2012

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Proteomic-based studies of nasal lavage fluid (NLF) may identify molecular pathways associated with disease pathology and new biomarker candidates of upper airway diseases. However, most studies have used rather tedious untargeted MS techniques. Selected reaction monitoring (SRM) is a sensitive and specific technique that can be used with high throughput. In this study, we developed a semiquantitative SRM-based method targeting 244 NLF proteins. The protein set was identified through a literature study in combination with untargeted LC-MS/MS analyses of trypsin-digested NLF samples. The SRM assays were designed using MS/MS data either downloaded from a proteomic data repository or experimentally obtained. Each protein is represented by one to five peptides, resulting in 708 SRM assays. Three to four transitions per assay were used to ensure analyte specificity. The majority (69%) of the assays showed good within-day precision (coefficient of variation ≤ 20%). The accuracy of the method was evaluated by analyzing four samples prepared with varying amounts of four proteins. Peptide and protein ratios were in good agreement with expected ratios. In conclusion, a high throughput screening method for relative quantification of 244 NLF proteins was developed. The method should be of general use in any proteomic study of the upper airways.

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“…LiCl in PBS was used to facilitate quantification of the exact amount of the middle ear effusions in the samples as described [24]. The lithium concentration was assayed by using the Vitros 250 analyser (Chemistry System, Johnson & Johnson Clinical Diagnostics, Rochester, NY, USA).…”

Section: Methodsmentioning

confidence: 99%

Cytolytic complement activity in otitis media with effusion

Närkiö-Mäkelä

Meri

2001

Clinical and Experimental Immunology

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SUMMARYOtitis media with effusion (OME) is a chronic inflammation persisting in the middle ear cavity of at least 8 weeks duration. Middle ear effusion (MEE; n 38), samples from children suffering from OME were investigated for their direct cytolytic activity or an ability to enhance complement lysis of unsensitized bystander cells. Thirteen of the 38 MEEs had direct endogenous haemolytic activity and 27 samples had an ability to enhance serum-initiated lysis. Using an enzyme immunoassay, high levels of terminal complement complexes (TCC) were detected in the MEE samples (mean 34´1 mg/ml, range 5±89 mg/ ml). This indicated strong local complement activation that had progressed to the terminal stage. As one potential factor promoting complement activation we identified both monomeric and trimeric properdin in MEE by Western blotting. By stabilizing C3 and C5 convertases properdin accelerates the alternative and terminal pathways of complement. On the other hand, the membrane attack complex (MAC) inhibitor CD59, which was found to be extensively shed into the MEE in a functionally active form, may control excessive cytotoxicity of the MEE. In conclusion, intense complement activation, up to the terminal level, maintains ongoing inflammation in the middle ear cavity and can pose a threat to the local epithelium.

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New method to assess dilution of secretions for immunological and microbiological assays

Cited by 11 publications

References 11 publications

Dilution assessment of cervicovaginal secretions obtained by vaginal washing for immunological assays

Dilution assessment of cervicovaginal secretions obtained by vaginal washing for immunological assays

Screening Method Using Selected Reaction Monitoring for Targeted Proteomics Studies of Nasal Lavage Fluid

Cytolytic complement activity in otitis media with effusion

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