New-onset afebrile seizures in infants: role of neuroimaging.

OBJECTIVE: To investigate the presenting characteristics of new-onset afebrile seizures in infants (age 1-24 months) and the yield of neuroimaging. METHODS: Prospective data were obtained from a standardized evaluation and management plan mandated by a critical care pathway. A total of 317 infants presented with new-onset afebrile seizures between 2001 and 2007. EEG was performed on 90.3%, head CT was obtained on 94%, and MRI was obtained on 57.4%. RESULTS: We found half of the infants had partial features to their seizures, yet evidence for primary generalized seizures was rare. The majority had more than 1 seizure upon presentation. Seizures in this age group tended to be brief, with 44% lasting less than 1 minute. EEG abnormalities were found in half. One-third of CTs were abnormal, with 9% of all CTs requiring acute medical management. Over half of MRIs were abnormal, with cerebral dysgenesis being the most common abnormality (p < 0.05). One-third of normal CTs had a subsequent abnormal MRI-only 1 resulted in altered medical management. CONCLUSIONS: Infantile seizures are usually brief, but commonly recurrent, and strong consideration should be made for inpatient observation. Acute imaging with CT can alter management in a small but important number of infants. Due to the superior yield, strong consideration for MRI should be given for all infants, as primary generalized seizures are rare, and there is a high rate of cerebral dysgenesis.

Engineering enzymes for clinical diagnosis.

The enzyme creatine amidinohydrolase (creatinase, EC 3.5.3.3.) is a major limiting factor in the enzymatic creatinine determination because of its comparatively poor catalytic activity and stability in the native form. The gene from Pseudomonas putida coding for creatinase was cloned and used for overexpression of the protein in E coli and Pseudomonas. In addition, it was possible by means of 'random' mutagenesis in vivo and subsequent screening using an activity plate assay to isolate creatinase derivatives that are more stable towards detergents and elevated temperature under test kit conditions. This example shows that enzymes can be optimized for use in given assay conditions by mutagenesis of cloned DNA and suitable screening methods.

Immunoturbidimetric method for routine determinations of apolipoproteins A-I, A-II, and B in normo- and hyperlipemic sera compared with immunonephelometry.

We describe a method for routine immunoturbidimetry of apolipoproteins (apo) A-I, A-II, and B in both normo- and hyperlipemic sera. A special antiserum reagent, consisting of a highly concentrated mixture of nonionic and anionic detergents (final concentration in the assay, 36 g/L), rapidly removes intrinsic turbidities of even strongly lipemic sera without interfering with the antigen-antibody precipitation reaction. The method has good precision, and obviates the need for special sample pretreatment, extended incubation periods, and measurment of sample blanks. A comparison with established immunoephelometric assays generally showed close agreement for analytical recoveries of the three apolipoproteins. However, in samples containing greater than or equal to 18 g of triglycerides per liter, the nephelometric assays yielded about two- to threefold higher values for apo A-II and B than did the turbidimetric procedure. To elucidate this discrepancy, we used the turbidimetric methods to assay sera with and without enzymatic lipolytic pretreatment. Even for samples with triglyceride concentrations up to 60 g/L, complete enzymatic lipolysis (as evidenced by thin-layer chromatography) did not significantly alter the recoveries of apo A-II and B from those obtained with the untreated specimens. Thus the immunoturbidimetric methods yield reliable results for apo A-I, A-II, and B, not only in normo- but also in hyperlipemic sera.

Multicentre evaluation of an enzymatic method for creatinine determination using a sensitive colour reagent.

A new enzymatic colorimetric method for the determination of creatinine in serum and urine (Creatinine PAP, Cat. No. 839434 and 836885, Boehringer Mannheim GmbH, Mannheim, FRG) was evaluated in 16 clinical chemistry laboratories and the manufacturer's testing laboratory. The test is based on the enzymatic degradation of creatinine and its reaction products by creatininase, creatinase and sarcosine oxidase. The H2O2 produced by the oxidation of sarcosine is determined using a modified Trinder reaction. The test can be carried out either manually or in mechanized analysers which enable the pipetting of a starter reagent to be made. The following results were obtained: Depending on the analyte concentration (range 40 to 1240 mumol/l), medians for the coefficients of variation were: 4.6-0.9% within-run and 6.4-2.8% between-day. At 546 nm the linear measuring range extended from 13 mumol/l (detection limit) to 1780 mumol/l, at 510 nm from 9 to 890 mumol/l. Recoveries in aqueous and human serum based standards as well as method comparisons with Fuller's earth methods and an enzymatic UV test indicate a high accuracy of this new enzymatic method in serum and urine. No interference was observed with haemolysed and lipaemic sera. This also applied to anticoagulants and to 36 drugs at therapeutic concentrations, with the exception of calcium dobesilate, which led to decreased values. Icteric samples containing 120-310 mumol/l bilirubin invariably led to decreased creatinine values (10-50 mumol/l lower). In a collaborative study substantially better interlaboratory agreement was observed with the new colorimetric enzymatic test than with the comparison methods (enzymatic UV test and various Jaffe procedures). In conclusion, this new enzymatic colorimetric test permits a precise and specific determination of creatinine in serum and urine. It makes a considerable contribution to improving the interlaboratory comparability of creatinine determinations and is suitable for routine use.

Reagent for the enzymatic determination of serum total cholesterol with improved lipolytic efficiency.

We describe a sensitive method for quantifying the extent of cholesterol ester cleavage during enzymatic assay of total cholesterol in serum. Lipids are extracted from the assay mixture with chloroform/methanol (1/1 by vol), concentrated, then quantified by "high-performance" thin-layer chromatography. Although with conventional enzymatic reagents for determination of serum total cholesterol the hydrolysis of the cholesterol esters may be incomplete, a new enzymatic cholesterol reagent (Monotest Cholesterol, High Performance, Boehringer Mannheim) gives virtually complete cholesterol ester cleavage (i.e., greater than or equal to 99.5%). Use of this reagent with its improved lipolytic efficiency yields results for serum total cholesterol that are identical to those measured with a candidate reference procedure involving alkaline cholesterol ester saponification.