Approximately one-quarter of circulating cobalamin (vitamin B-12) binds to transcobalamin (holoTC) and is thereby available for the cells of the body. For this reason, holoTC is also referred to as active vitamin B-12. HoloTC was suggested as an optimal marker of early vitamin B-12 deficiency >20 y ago. This suggestion led to the development of suitable assays for measurement of the compound and clinical studies that aimed to show the benefit of measurement of holoTC rather than of vitamin B-12. Today holoTC can be analyzed by 3 methods: direct measurement of the complex between transcobalamin and vitamin B-12, measurement of vitamin B-12 attached to transcobalamin, or measurement of the amount of transcobalamin saturated with vitamin B-12. These 3 methods give similar results, but direct measurement of holoTC complex is preferable in the clinical setting from a practical point of view. HoloTC measurement has proven useful for the identification of the few patients who suffer from transcobalamin deficiency. In addition, holoTC is part of the CobaSorb test and therefore useful for assessment of vitamin B-12 absorption. Clinical studies that compare the ability of holoTC and vitamin B-12 to identify individuals with vitamin B-12 deficiency (elevated concentration of methylmalonic acid) suggest that holoTC performs better than total vitamin B-12. To date, holoTC has not been used for population-based assessments of vitamin B-12 status, but we suggest that holoTC is a better marker than total vitamin B-12 for such studies.
The high cobalamin levels were mainly explained by high levels of holoTC, possibly caused by complex formation with its soluble receptor, sCD320. The family occurrence points to a genetic explanation.
Transcobalamin (TC) deficiency (OMIM# 275350) is a rare, autosomal recessive disorder that presents in early infancy with a broad spectrum of symptoms, including failure to thrive, megaloblastic anemia, immunological deficiency, and neurological symptoms. Here we report a study of a family (parents and three children) with two children suffering from TC deficiency caused by two different mutations in the TCN2 gene. Initially, molecular genetic analysis of genomic DNA revealed a heterozygous mutation in the +1 position of exon 7 (c.1106+1 G > A) in the father and all three children. Bioinformatic analysis indicates that this mutation causes exon skipping, and further experiments supported this hypothesis and suggested that the mutant allele undergoes nonsense-mediated messenger RNA (mRNA) decay. We did not identify further mutations in genomic DNA that could explain TC deficiency in the two children. However, further efforts using complementary DNA (cDNA) derived from RNA from blood leukocytes identified a large deletion removing the entire exon 8, resulting in a frameshift and a premature stop codon (p.E371fsX372) in the mother and the two affected children. Our data indicate that if exon-by-exon DNA sequencing of genomic DNA does not uncover mutations corresponding to the phenotype, a systematic search for other mutations should be initiated by sequencing cDNA or using semiquantitative methods to detect large deletions in TCN2.
In Denmark, biennial population screening for colorectal cancer was introduced in 2014 for all aged 50-74 years. Five laboratories representative for the regional division of Denmark perform the immunochemical testing of faecal occult blood in the screening samples (iFOBT, OC-Sensor (Eiken Chemical, cut-off 100 µg/L)). In July 2016, a new agreement on the public post-delivery entailed an increased lag time (five days) from the screening participant drops the screening sample into a mail-box until sample arrival at the laboratories. Previous work had reported that a lag time above five days led to more false negative iFOBT tests. We investigated if this was true also under Danish conditions. We performed two stability tests; one with sample storage at 30 °C for 14 days (N = 60), and another with sample storage at room temperature for 13 days (N = 10). We extracted data from our laboratory information system (LABKA) on all iFOBT tests performed in the entire Central Denmark Region (N = 104,328 patients) during the last six months for each calendar year 2014-16. For each year, we computed the distribution of iFOBT tests below and above cut-off. Our stability tests showed no positive samples switching to false negative after storage; however, some negative samples turned false positive, especially at 30 °C. The data showed no change in the distribution of iFOBT tests below and above cut-off after July 2016. We found no evidence that an enhanced lag time increased the number of false negative iFOBT tests in the Danish screening program for colorectal cancer.
Artificial sweeteners are widely used as substitutes for sugar. The sweeteners are generally considered safe, however their whereabouts during pregnancy and lactation and the effect on child development are poorly explored. There is a need for new tools to measure these substances during pregnancy and lactation. Here, we describe the development and validation of a sensitive liquid chromatographytandem mass spectrometry method for the simultaneous quantification of acesulfame, cyclamate, saccharin and sucralose in human plasma, umbilical cord blood, amniotic fluid and breast milk. The samples were prepared by protein precipitation and separated on a Luna Omega Polar C 18 column (2.1 Â 50 mm, 1.6 μm).Electrospray ionization in negative mode and multiple reaction monitoring were used to monitor the ion transitions. The validated concentration ranges were from 1 to 500 ng/ml (10-500 ng/ml for sucralose). Interassay precisions were all ≤15% and the accuracies were within ±15%. Stability, linearity, dilution integrity, carryover and recovery were also examined and satisfied the validation criteria. Finally, this analytical method was successfully applied on spiked samples of plasma, umbilical cord blood, amniotic fluid and breast milk, proving its suitability for use in clinical studies on artificial sweeteners, including during pregnancy and lactation.
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