Loss of human chorionic gonadotropin in urine during storage at − 20 °C

Lempiäinen, Anna; Hotakainen, Kristina; Alfthan, Henrik; Stenman, Ulf‐Håkan

doi:10.1016/j.cca.2011.09.038

Cited by 28 publications

(17 citation statements)

References 27 publications

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“…For example, prestorage extraction by acetone precipitation [12,13] or by addition of glycerol or albumin [13][14][15] stabilize immunoreactivity. [19] This is consistent with the stability of a urinary LH assay based on a different strategy of pre-assay dissociation into subunits with detection aimed at the LH beta subunit, [20] an assay displaying prolonged stability during frozen storage. [19] This is consistent with the stability of a urinary LH assay based on a different strategy of pre-assay dissociation into subunits with detection aimed at the LH beta subunit, [20] an assay displaying prolonged stability during frozen storage.…”

Section: Introductionsupporting

confidence: 70%

“…[13,29] Validation after prolonged frozen conditions has not been evaluated by available commercial immunoassays. [17,37,38] Loss in urinary LH and hCG immunoreactivity after 7 to 70 days of storage at -20 C (unpreserved) measured with the IF immunoassay, [13,19] radioimmunoassay, [14] and immunoenzymatic assay [27] was also reported previously which was attributed to the dissociation of the heterodimeric glycoprotein into its sub-units in the presence of urinary urea in specimens. [30][31][32] This diversity reflects the metabolism of the gonadotropins which produces nicked dimers and free sub-units including the core fragment.…”

Section: Discussionmentioning

confidence: 70%

“…These discrepancies between LH immunoassays most probably reflect the diversity of immunoreactive LH species in human urine compared with the bloodstream. [14,19] Loss of immunoreactivity of urinary LH (measured with IF immunoassay) was also reported after 24 weeks of -80 C storage. Like hCG where the metabolism is more fully characterized, [33] the homologous LH heterodimer is metabolized and excreted in the urine in various fragments which are largely in the forms of free sub-units and fragmented forms of LHb.…”

Section: Discussionmentioning

confidence: 97%

“…[11] Urine LH immunoreactivity may depend on urine sample storage conditions as well as immunoassay epitope specificity. Furthermore, whereas two-site assays with epitopes on different subunit of the heterodimer optimizes detection of intact LH, [16][17][18] dissociation of LH into its sub-units, influenced by exposure to high urinary concentrations of urea, [19] may be reflected in loss of immunoreactivity whether or not the subunit remain intact especially at -20 C storage. Furthermore, whereas two-site assays with epitopes on different subunit of the heterodimer optimizes detection of intact LH, [16][17][18] dissociation of LH into its sub-units, influenced by exposure to high urinary concentrations of urea, [19] may be reflected in loss of immunoreactivity whether or not the subunit remain intact especially at -20 C storage.…”

Section: Introductionmentioning

confidence: 99%

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Immunoreactive LH in long‐term frozen human urine samples

Singh

Jimenez

Newman

et al. 2013

Drug Testing and Analysis

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Urine provides a convenient non-invasive alternative to blood sampling for measurement of certain hormones. Urinary luteinizing hormone (LH) measurements have been used for endocrinology research and anti-doping testing. However, the commercially available LH immunoassays are developed and validated for human blood samples but not urine so that LH assays intended for use with urine samples need thorough validation. Therefore, the present study evaluated the measurement of urinary LH immunoreactivity using previously validated immunofluorometric (IF) and immunochemiluminometric (ICL) LH assays after prolonged frozen storage. LH was measured in serial urine samples following administration of a single injection of one of two doses of recombinant human chorionic hormone (rhCG) with assays run at the end of study (2008) and again after four years of frozen (-20 °C) storage where samples were stored without adding preservatives. The ICL assay showed quantitatively reproducible LH measurements after prolonged -20 °C storage. However, the IF immunoassay gave consistently lower LH levels relative to ICL (2008) with a further proportionate reduction after four years of sample storage (2012). Yet, both the assays displayed similar patterns of the time-course of urine LH measurement both before and after four years of frozen storage. In conclusion, we found that both immunoassays are suitable for urinary LH measurements with ICL assay being more robust for quantitative urinary LH measurement such as for anti-doping purposes, whereas the IF could be applicable for research studies where urine LH levels are compared within-study but not in absolute terms.

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Section: Introductionsupporting

confidence: 70%

Section: Discussionmentioning

confidence: 70%

Section: Discussionmentioning

confidence: 97%

Section: Introductionmentioning

confidence: 99%

See 2 more Smart Citations

Immunoreactive LH in long‐term frozen human urine samples

Singh

Jimenez

Newman

et al. 2013

Drug Testing and Analysis

View full text Add to dashboard Cite

show abstract

“…Comparison of the MS method with an immunochemistry‐based reference method currently used in WADA‐accredited doping laboratories showed strong correlation in serum (Lund et al, ). In urine, on the other hand, the MS method reported significantly higher hCG levels than the immunoassay; probably due to molecular dissociation of intact hCG‐heterodimer into free subunits during urine storage (Lempiainen et al, ). Because the MS method quantified both intact hCG and free hCG‐β, it was less susceptible to this problem and demonstrated another benefit of more wide‐spread implementation of MS strategies.…”

Section: Bottom‐up Ms Detection Of Growth‐promoting Peptides and Protmentioning

confidence: 99%

Current trends in mass spectrometry of peptides and proteins: Application to veterinary and sports‐doping control

Broek

Blokland

Nessen

et al. 2013

Mass Spectrometry Reviews

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Detection of misuse of peptides and proteins as growth promoters is a major issue for sport and food regulatory agencies. The limitations of current analytical detection strategies for this class of compounds, in combination with their efficacy in growth-promoting effects, make peptide and protein drugs highly susceptible to abuse by either athletes or farmers who seek for products to illicitly enhance muscle growth. Mass spectrometry (MS) for qualitative analysis of peptides and proteins is well-established, particularly due to tremendous efforts in the proteomics community. Similarly, due to advancements in targeted proteomic strategies and the rapid growth of protein-based biopharmaceuticals, MS for quantitative analysis of peptides and proteins is becoming more widely accepted. These continuous advances in MS instrumentation and MS-based methodologies offer enormous opportunities for detection and confirmation of peptides and proteins. Therefore, MS seems to be the method of choice to improve the qualitative and quantitative analysis of peptide and proteins with growth-promoting properties. This review aims to address the opportunities of MS for peptide and protein analysis in veterinary control and sports-doping control with a particular focus on detection of illicit growth promotion. An overview of potential peptide and protein targets, including their amino acid sequence characteristics and current MS-based detection strategies is, therefore, provided. Furthermore, improvements of current and new detection strategies with state-of-the-art MS instrumentation are discussed for qualitative and quantitative approaches.

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Annual banned‐substance review: analytical approaches in human sports drug testing

Thevis¹,

Kuuranne

Geyer

et al. 2012

Drug Testing and Analysis

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Frequently updated anti-doping regulations and detection strategies for banned substances and methods of doping are vital for efficient and scientifically substantiated abatement of drug abuse, manipulation, and illicit performance enhancement in sport. In 2012, the ninth version of the Prohibited List as annually issued by the World Anti-Doping Agency (WADA) reports on modest but relevant modifications from its 2011 predecessor. Awareness of old and new analytical challenges in sports drug testing, together with issues resulting from altered rules and regulations, have given rise to numerous research projects aiming at improved detection strategies to strengthen international anti-doping efforts. In this annual banned substance review, emerging and advancing methods in the detection of known and recently outlawed substances are reported. New and/or enhanced procedures and techniques of doping analysis are reviewed together with information relevant to doping controls. The review surveys and critiques literature published between October 2011 and September 2012.

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Loss of human chorionic gonadotropin in urine during storage at − 20 °C

Cited by 28 publications

References 27 publications

Immunoreactive LH in long‐term frozen human urine samples

Immunoreactive LH in long‐term frozen human urine samples

Current trends in mass spectrometry of peptides and proteins: Application to veterinary and sports‐doping control

Annual banned‐substance review: analytical approaches in human sports drug testing

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