Abstract C-reactive protein (CRP) is able to bind phosphocholine in the presence of calcium ions. According to a previous functional property of CRP, we tried to develop an affordable and cheap high sensitive nephelometric CRP assay using soy oil. Serum samples were measured by Nephelometer BNII (Siemens), by mixing the serum with diluted soy oil emulsion (Intralipid ® 20%) and Tris-calcium buffer (PH 7.5). The measurement took place after 12 min incubation time at 37°C by measuring the agglutination between CRP and phosphocholine. Results from our automated functional assay were compared with results obtained using an immunoturbidimetric CRP assay. Results showed a good correlation coefficient for method comparison between functional nephelometric CRP assay and immunoturbidimetric CRP assay, r = 0.895, significance level p <0.0001. The limit of detection for the functional nephelometric CRP assay was 0.1 mg/L. However, the within run % CV values for the functional assay were 6.1 % (20 mg/L), 4.7 % (50 mg/L) and 4.5 % (100 mg/L). The between-run % CV values were 17.6 % (20 mg/L), 18.8 % (50 mg/L), and 11.3 % (100 mg/L). The new functional nephelometric CRP assay enables high sensitive CRP measurement in serum in the range of 0.1 mg/L to 300 mg/L. The functional assay could be used for veterinary analysis due to the ability to measure CRP according to the functional properties, not the morphological properties which depend on specific antibodies.

Hematology analyzers can be used for screening patients for blood abnor-malities. The techniques used in a hematology analyzer include impedanceanalysis, flow cytometry and spectroscopy, which allow for measuring of forexample absolute count, sizes and concentration of different cells in a patient’sblood sample. Hyperlipidemia, which refers to elevated blood lipid levels, isthe primary cause of heart-related illness and fatalities in today’s developedor developing countries. Currently, blood lipid levels are not measured as aparameter with hematology analyzers. Since hematology analyzers allow for arapid general screening of blood parameters, an area of interest is therefore tobe able to measure blood lipids with a hematology analyzer. Thus, this studyaims to investigate the possibility of detecting and measuring blood lipids witha hematology analyzer, using flow cytometry and/or spectrophotometry. In order to investigate this possibility, two simulating methods were conductedwhere in the first method Intralipid 20% was mixed with saline into sampleswith different lipid concentrations. In the second method, diluent wasused instead of saline. Lastly a Correlation study was performed whereIntralipid 20% was mixed with donor blood to prepare samples with differentlipid concentrations. All samples were then analyzed in a hematologyanalyser and scatter plots from flow cytometry and light absorption datafrom spectrophotometry measurements were obtained. The methods showedthat there is a strong correlation between number of detected pulse countsfrom the scatter plots and lipid concentration. Same applies to lightabsorption compared to the lipid concentration of the samples, measured withspectrophotometry. The results from this study show that it is in fact possible to detect andmeasure blood lipid levels with a hematology analyser using flow cytometryand spectrophotometry. Further development within this area could thereforeenable simple screening of this additional parameter and early detection ofindications of hyperlipidemia. / Hematologianalysatorer möjliggör screening av eventuella avvikelser ipatienters blood. De tekniker som används i en hematologianalysatorinkluderar impedansanalys, flödescytometri och spektroskopi, vilka möj-liggör mätning av till exempel absolutantal, storlekar och koncentrationav olika celler i ett patientblodprov. Hyperlipidemi, vilket hänvisar tillförhöjda blodlipidnivåer, är den främsta orsaken till hjärtrelaterade sjukdomaroch dödsfall i dagens utvecklade eller utvecklingsländer. För närvarandemäts inte blodlipidnivåer som en parameter med hematologianalysatorer.Eftersom hematologianalysatorer möjliggör en snabb allmän screening avblodparametrar, är ett intresseområde därför att kunna mäta blodlipider meden hematologianalysator. Syftet med denna studie är därför att undersökamöjligheten att detektera och mäta blodlipider med en hematologianalysator,med hjälp av flödescytometri och/eller spektrofotometri. Två simuleringsmetoder genomfördes, där den första metoden innefattadeblandning av Intralipid 20% med saltlösning till prover med varierandelipidkoncentration. I den andra metoden användes spädningsvätska iställetför saltlösning. Slutligen genomfördes en korrelationsstudie där Intralipid20% blandades med donatorblod och prover med olika lipidkoncentrationerförbereddes. Alla prover analyserades sedan i en hematologianalysator ochspridningsdiagram och ljusabsorptionsdata från spektrofotometrimätningarerhölls. Resultaten visade att det finns en stark korrelation mellan antalet de-tekterade pulsräkningar från spridningsdiagrammen och lipidkoncentrationen.Samma gäller för ljusabsorption jämfört med lipidkoncentrationen i proverna,mätt med spektrofotometri. Resultaten från denna studie visar att det faktiskt är möjligt att detektera ochmäta blodlipidnivåer med en hematologianalysator med hjälp av flödescyto-metri och spektrofotometri. Vidare utveckling inom detta område skulle därförkunna möjliggöra enkel screening av patienters blod lipidkoncentration samtunderlätta en tidig upptäckt av indikationer på hyperlipidemi.

Introduction: Post-mortem whole blood samples differ greatly in quality, lipemia is one cause of concern in toxicological analyses. Around 4 % of all samples sent to RMV are given a notation of lipemic content. The aim of the thesis was to study the effects of lipemia on the quantification of 14 benzodiazepines and 5 similar sedative and antianxiety drugs as well as evaluate the pre-analytical process aiming to reduce the effects of lipemia.  Methods: Blood samples were simulated with bovine blood, analyte spiking, and lipid spiking with either the nutrition emulsion Intralipid or with a mixture of post-mortem lipids from authentic samples. The outset was the by RMV currently used LLE method followed by UPLC- MS/MS and the extraction method was altered and evaluated. Matrix effects were also studied.  Results: Lipemia were found to be a great interference when quantifying benzodiazepines. For most analytes, internal standard could compensate for the loss of analyte but there was a problem with analytes not having their own IS. The 7-amino-compounds were greatly affected by lipemia and propiomazine and dihydropropiomazine showed extreme losses. Equilibration of IS did not result in similar loss as analyte. Dilution of sample reduced losses caused by lipemic content. SPE resulted in extracts free from lipids and high yields but there were analyte losses similar to LLE. No matrix effects from the lipids were found. Samples spiked with Intralipid gave poorer analyte yields than those spiked with post-mortem lipids.  Conclusion: Dilution is the most successful method to reduce pre-analytical matrix effects as long as the concentration is not so low that it risks getting lower than the analytical limits when doing so. Not homogenising samples before sampling is giving incorrect results. SPE could, if optimised for the analyte retention and elution, remove lipids from samples and obtain accurate analyte concentrations. Pooling lipids from post-mortem samples is a possible method for simulating lipemic whole blood. Intralipid and the PM-mix gave the same indications, but to different extents. Further studies where the ability to mimic authentic lipids are needed for both Intralipid and PM-mix.

<p>To estimate the propagation of light in tissue-like optical phantoms (fat emulsions), this thesis utilized the diffuse reflectance spectroscopy in combination with Monte Carlo simulations. A method for determining the two-parametric Gegenbauer-kernal phase function was utilized in order to accurately describe the diffuse reflectance from poly-dispersive scattering optical phantoms with small source-detector separations. The method includes the spectral collimated transmission, spatially resolved diffuse reflectance spectra (SRDR) and the inverse technique of matching spectra from Monte Carlo simulations to those measured. An absolute calibration method using polystyrene micro-spheres was utilized to estimate the relation between simulated and measured SRDR intensities. The phase function parameters were comparable with previous studies and were able to model measured spectra with good accuracy. Significant differences between the phase functions for homogenized milk and the nutritive fat emulsions were found.</p><p> </p>

To estimate the propagation of light in tissue-like optical phantoms (fat emulsions), this thesis utilized the diffuse reflectance spectroscopy in combination with Monte Carlo simulations. A method for determining the two-parametric Gegenbauer-kernal phase function was utilized in order to accurately describe the diffuse reflectance from poly-dispersive scattering optical phantoms with small source-detector separations. The method includes the spectral collimated transmission, spatially resolved diffuse reflectance spectra (SRDR) and the inverse technique of matching spectra from Monte Carlo simulations to those measured. An absolute calibration method using polystyrene micro-spheres was utilized to estimate the relation between simulated and measured SRDR intensities. The phase function parameters were comparable with previous studies and were able to model measured spectra with good accuracy. Significant differences between the phase functions for homogenized milk and the nutritive fat emulsions were found.