Procedures For Msms Screening

Specimen preparation for MS-MS screening requires an extraction and derivatization process,[3] which is performed using 96-well polypropylene microplates in routine analysis.[4-6] One 3.2-mm disk of dried blood spot is punched out into the individual wells of a microplate. To the disk in each well, a methanol solution containing known concentrations of stable isotope-labeled standards is added. The microplate is covered and the samples are mixed. The extract is transferred to a second microplate and dried. The residue in each well is derivatized with butanolic HCl. After removing the excess reagent in each well, the derivatized residue is dissolved in an acetonitrile-water mixture. Then the microplate is sealed and placed in the autoinjector tray for ESI-MS-MS analysis.

In routine analysis under the control of the computer system, intensities of designated ionic compounds of the sample are measured in multiple modes of analysis, and a set of data for the sample is recorded sequentially. The amounts of the compounds as diagnostic markers are calculated using the assumed volume of blood in a punch and the intensities of both the compounds and the respective stable isotope-labeled internal standards. At present, not all internal standards for acylcarnitine analysis are available, and the amounts of some acylcarnitines are calculated using the labeled acylcarnitine with different acyl groups (e.g., deuterium-labeled C14-acylcarnitine for C14:1-acylcarnitine measurement and deuterium-labeled C16-acylcarnitine for hydroxy-C16-acylcarnitine measurement).

For samples with abnormal values for any diagnostic marker, repunched blood spots are analyzed to confirm the initial screening. Then resamplings for newborns with abnormal values are performed to obtain additional clues to the suspected disease and to confirm that the correct

Target diseases in MS-MS screening are listed in Table 1.

The clinical severity of target diseases, except for PKU and homocystinuria (HCU), is quite heterogeneous.[7,8] Patients with severe forms of the diseases present clinical crises with metabolic acidosis, hyperammonemia, and/or hypoglycemia in the newborn period and may not survive in spite of intensive care. Liver transplantations have been tried for patients with severe forms of organic acidemias and urea cycle disorders. Patients with milder forms may experience intermittent and sometimes life-threatening crises because of catabolic conditions with or without progressive brain dysfunction. Other symptoms of fatty acid oxidation disorders are muscle weakness and pain with high CK values. Some patients with mild forms of the diseases may not show any symptoms without medical intervention.

Long-term therapies for most diseases include low-protein and/or low-fat diets. Depending on the respective diseases, carnitine supplementation, coenzyme administration, avoidance of fasting, and treatment of acute episodes with intravenous infusion of glucose may be required. Medium-chain triglycerides are used for some fatty acid oxidation disorders, and sodium benzoate or alternatives are for hyperammonemia of urea cycle disorders.

Diagnostic Markers for Target Diseases

The cutoffs for diagnostic markers (Table 1) are set empirically in most laboratories based on the data from the retrospective analysis of the patients and the prospective studies. High false-positive rates have been reported in newborns with very low birth weights or those requiring intensive care.[6] To improve the sensitivity for screening, additional markers, such as the ratio of the main marker to the second marker, are proposed.

High levels of free carnitine and low levels of long-chain acylcarnitines in blood spots characterize carnitine

Fig. 2 Fragmentation of amino acid butyl esters in a collision chamber.

Fig. 2 Fragmentation of amino acid butyl esters in a collision chamber.

Getting Started With Dumbbells

Getting Started With Dumbbells

The use of dumbbells gives you a much more comprehensive strengthening effect because the workout engages your stabilizer muscles, in addition to the muscle you may be pin-pointing. Without all of the belts and artificial stabilizers of a machine, you also engage your core muscles, which are your body's natural stabilizers.

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