Elevated concentration of C-reactive protein is associated with pregnancy-related co-morbidities but not with relapse activity in multiple sclerosis.

During pregnancy, alterations take place in mother's immune system with the goal of maintaining a successful pregnancy, and delivering healthy offspring. Immune alterations include activation of the innate immune system and dampening of cell-mediated adaptive immunity. Due to these alterations, cell-mediated autoimmune diseases typically ameliorate during pregnancy. The objectives of this study were to evaluate whether C-reactive protein (CRP) concentration, a sensitive marker of systemic inflammation (1) is increased during MS pregnancy (2) predicts pregnancy-related co-morbidities associated with MS (3) predicts MS disease activity after delivery. CRP concentration was measured using a high sensitivity assay from seven prospectively collected serum samples of 41 MS patients and 19 controls during pregnancy and 6 months after delivery. Annualized relapse rates, EDSS, fatigue scores and obstetric details of the patients were recorded. Delivery-related CRP levels were significantly elevated both among MS patients and in controls. CRP levels were higher during pregnancy than during the postpartum period in both study groups. Delivery-related elevated CRP levels did not correlate with postpartum disease activity. MS patients with eventual gestational diabetes had a significantly higher median CRP in the beginning of pregnancy compared to non-diabetic MS patients (9.28 vs. 2.98 mg/l, p = 0.0025). MS patients reporting fatigue had a significantly higher CRP throughout pregnancy compared to patients without fatigue. Higher CRP values were associated with pregnancy-related co-morbidities but not with MS disease activity.

Adaptation of the human skin by chronic solar-simulating UV irradiation prevents ultraviolet-B irradiation-induced rise in serum C-reactive protein levels.

Exposure of the skin to UV radiation induces local inflammation. We hypothesized that inflammation induced by erythemal UV-B irradiation could elevate levels of serum C-reactive protein (CRP) and that suberythemal repeating doses of solar-simulating UV radiation (SSR) would produce photoadaptation to such inflammation. Separation-free high-sensitivity assays of CRP show an increase by 42% (P = 0.046) in CRP concentrations in healthy human subjects 24 h after a 3 minimal erythemal dose (MED) dose of UV-B delivered onto a 100 cm2 skin area. Preceding daily suberythemal doses of whole-body SSR for 10 or 30 consecutive days completely prevented the CRP increase. UV-B-induced skin erythema was partially attenuated by 30 preceding days of SSR only (P = 0.00066). After 10 daily SSR doses, the mean baseline CRP concentrations (0.24 +/- 0.21 mg/L) declined by 35% (P = 0.018). Using high-sensitivity analysis of serum CRP as the endpoint marker for cutaneous inflammation, we show that acute exposure of even a relatively small skin area to erythemal UV-B induces skin inflammation detectable also at the systemic level and that photoadaptation by preceding repeating suberythemal doses of SSR reduces signs of inflammation. Our data complement the view given by previous studies in that local photoadaptation also has systemic manifestations.

Fluorescent nanoparticles as labels for immunometric assay of C-reactive protein using two-photon excitation assay technology.

We describe the use of fluorophore-doped nanoparticles as reporters in a recently developed ArcDia TPX bioaffinity assay technique. The ArcDia TPX technique is based on the use of polymer microspheres as solid-phase reaction carrier, fluorescent bioaffinity reagents, and detection of two-photon excited fluorescence. This new assay technique enables multiplexed, separation-free bioaffinity assays from microvolumes with high sensitivity. As a model analyte we chose C-reactive protein (CRP). The assay of CRP was optimized for assessment of CRP baseline levels using a nanoparticulate fluorescent reporter, 75 nm in diameter, and the assay performance was compared to that of CRP assay based on a molecular reporter of the same fluorophore core. The results show that using fluorescent nanoparticles as the reporter provides two orders of magnitude better sensitivity (87 fM) than using the molecular label, while no difference between precision profiles of the different assay types was found. The new assay method was applied for assessment of baseline levels of CRP in sera of apparently healthy individuals.

Detection methods of microsphere based single-step bioaffinity and in vitro diagnostics assays.

Microspheres provide a solid phase substrate for bioaffinity binding similar to the walls of traditional test tubes and the wells of microtiter plates. The coated microsphere concentrates analyte molecules in the reaction volume on its surface. When the bioaffinity binding reaction has reached an equilibrium, the local concentration of the analyte in close proximity of the microsphere is orders of magnitude higher than the concentration of the analyte in the total reaction volume. The preparation and quality control of microspheres coated with bioactive material is less costly and labour intensive when compared to test tube or microwell plate coating procedures. In addition, the cost for logistics and transportation of microsphere reagents is lower than that of coated tubes or plates. Moreover, microspheres can be easily used in miniaturised assay formats and several different detection schemes can be employed in the measurement of microsphere-based assays. Several different types of microspheres are commercially available. The microspheres can be manufactured in different sizes from many materials, such as polystyrene, acrylate, and glass. The surface of the microspheres can be activated to enable covalent binding of biomolecules. Further, the microspheres may contain internal fluorochrome or magnetic material, for identification or separation purposes. In this paper we review different assay formats for single-step measurement of bioaffinity assays employing microspheres. The term single-step is used to describe assays where all reagents and the sample are mixed, incubated and measured without separate washing steps.

Two-photon excitation fluorometric measurement of homogeneous microparticle immunoassay for C-reactive protein.

Recent developments in infrared laser technology have enabled the design of a compact instrumentation for two-photon excitation microparticle fluorometry (TPX). The microparticles can be used in immunoassays as the antibody-coated solid phase to capture an antigen and then detect it with a fluorescently labeled tracer antibody. Unlike most other methods, TPX technology allows low-volume, homogeneous immunoassays with real-time measurements of assay particles in the presence of a moderate excess of fluorescent tracer. In this study, the TPX assay system was used for the reagent characterization and the measurement of C-reactive protein (CRP) in diluted plasma samples, targeting the assay range useful in infectious disease diagnosis. The pentameric structure of the CRP permitted the optimization of an assay with the lowest detectable concentration of 1 microg/L (7.5 pM) by using a single monoclonal antibody both for capture and as the tracer. With a 1:200 predilution of samples, the measurement range of the assay was 1-150 mg/L, but an additional 1:10 dilution was required for higher concentrations. The TPX method showed a good correlation with the reference result obtained in a routine hospital laboratory, demonstrating the feasibility of the technology for immunodiagnostic applications.