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Comparison of liquid chromatography-tandem mass spectrometry and sandwich ELISA for determination of keratan sulfate in plasma and urine.

Hintze JP, Tomatsu S, Fujii T, Montaño AM, Yamaguchi S, Suzuki Y, Fukushi M, Ishimaru T, Orii T - Biomark Insights (2011)

Bottom Line: KS is synthesized mainly in cartilage and released into circulation, making it a critical biomarker for MPS IVA to evaluate clinical course and effectiveness of therapies.No correlation was found between plasma KS measurements in controls.A moderate correlation between blood and urine KS measurements in the same individual was observed.

View Article: PubMed Central - PubMed

Affiliation: Department of Pediatrics, School of Medicine, Saint Louis University, St. Louis, MO, USA.

ABSTRACT

Background and aim: Mucopolysaccharidosis IVA (MPS IVA) leads to skeletal dysplasia through excessive storage of chondroitin-6-sulfate and keratan sulfate (KS). KS is synthesized mainly in cartilage and released into circulation, making it a critical biomarker for MPS IVA to evaluate clinical course and effectiveness of therapies. Therefore, an accurate and sensitive method is required to measure KS levels.

Material and methods: Using sandwich ELISA and liquid chromatography tandem mass spectrometry (LC/MS/MS) assays, we measured KS levels in blood and urine from MPS IVA patients and healthy controls to evaluate comparability of results. Blood (patients, n = 110; controls, n = 364) and urine (patients, n = 103; controls, n = 326) specimens were obtained.

Results: Plasma and urine KS measurements in patients were age-dependent and higher than age-matched controls. We observed a moderate correlation (r = 0.666; P < 0.001) between urine KS measurements and a weak correlation (r = 0.333; P = 0.002) between plasma KS measurements by ELISA and LC/MS/MS methods in patients. No correlation was found between plasma KS measurements in controls. The difference between KS measurements assayed by LC/MS/MS and ELISA was greater in controls than in patients. A moderate correlation between blood and urine KS measurements in the same individual was observed.

Conclusion: These findings indicate that both methods to measure blood and urine KS are suitable for diagnosis, monitoring therapies, and longitudinal assessment of the disease course in MPS IVA, but the LC/MS/MS method measures over 10 times more KS present in body fluids.

No MeSH data available.


Related in: MedlinePlus

Concentrations of plasma and urine KS of patients with MPS IVA and normal individuals as measured by ELISA and LC/MS/MS. A) Plasma KS results by ELISA of 109 specimens from MPS IVA individuals and 212 normal individuals are plotted with respect to age. B) Urine KS results by ELISA of 100 specimens from MPS IVA individuals and 179 normal individuals are plotted with respect to age. C) Plasma KS results by LC/MS/MS of 85 specimens from MPS IVA individuals and 165 normal individuals are plotted with respect to age. D) Urine KS results by LC/MS/MS of 58 specimens from MPS IVA individuals and 134 normal individuals are plotted with respect to age.
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f2-bmi-6-2011-069: Concentrations of plasma and urine KS of patients with MPS IVA and normal individuals as measured by ELISA and LC/MS/MS. A) Plasma KS results by ELISA of 109 specimens from MPS IVA individuals and 212 normal individuals are plotted with respect to age. B) Urine KS results by ELISA of 100 specimens from MPS IVA individuals and 179 normal individuals are plotted with respect to age. C) Plasma KS results by LC/MS/MS of 85 specimens from MPS IVA individuals and 165 normal individuals are plotted with respect to age. D) Urine KS results by LC/MS/MS of 58 specimens from MPS IVA individuals and 134 normal individuals are plotted with respect to age.

Mentions: We found that optimized KS calibration curves gave a good standard curve (R2 = 0.99) between 2.5 ng/mL and 40 ng/mL in ELISA (Fig. 1A) and 0.2 μg/ml and 10 μg/ml in LC/MS/MS (Fig. 1B). Plasma and urine KS measurements in MPS IVA patients were age-dependent and appreciably higher than age-matched controls (Tables 1 and 2; Figs. 2 and 3). Plasma KS concentrations measured by ELISA increased until age 10 years, at which point they fell gradually until normalizing around age 20 years. Plasma KS measurements by LC/MS/MS were highest in the first 5 years of life, and declined gradually until reaching a plateau near 20 years of age. Urine KS concentrations were highest during the first 5 years of life. After 5 years of age, urine KS levels declined until stabilizing around 20 years of age. This age-dependent pattern was observed with both ELISA and LC/MS/MS urine KS measurements.


Comparison of liquid chromatography-tandem mass spectrometry and sandwich ELISA for determination of keratan sulfate in plasma and urine.

Hintze JP, Tomatsu S, Fujii T, Montaño AM, Yamaguchi S, Suzuki Y, Fukushi M, Ishimaru T, Orii T - Biomark Insights (2011)

Concentrations of plasma and urine KS of patients with MPS IVA and normal individuals as measured by ELISA and LC/MS/MS. A) Plasma KS results by ELISA of 109 specimens from MPS IVA individuals and 212 normal individuals are plotted with respect to age. B) Urine KS results by ELISA of 100 specimens from MPS IVA individuals and 179 normal individuals are plotted with respect to age. C) Plasma KS results by LC/MS/MS of 85 specimens from MPS IVA individuals and 165 normal individuals are plotted with respect to age. D) Urine KS results by LC/MS/MS of 58 specimens from MPS IVA individuals and 134 normal individuals are plotted with respect to age.
© Copyright Policy - open-access
Related In: Results  -  Collection

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getmorefigures.php?uid=PMC3140273&req=5

f2-bmi-6-2011-069: Concentrations of plasma and urine KS of patients with MPS IVA and normal individuals as measured by ELISA and LC/MS/MS. A) Plasma KS results by ELISA of 109 specimens from MPS IVA individuals and 212 normal individuals are plotted with respect to age. B) Urine KS results by ELISA of 100 specimens from MPS IVA individuals and 179 normal individuals are plotted with respect to age. C) Plasma KS results by LC/MS/MS of 85 specimens from MPS IVA individuals and 165 normal individuals are plotted with respect to age. D) Urine KS results by LC/MS/MS of 58 specimens from MPS IVA individuals and 134 normal individuals are plotted with respect to age.
Mentions: We found that optimized KS calibration curves gave a good standard curve (R2 = 0.99) between 2.5 ng/mL and 40 ng/mL in ELISA (Fig. 1A) and 0.2 μg/ml and 10 μg/ml in LC/MS/MS (Fig. 1B). Plasma and urine KS measurements in MPS IVA patients were age-dependent and appreciably higher than age-matched controls (Tables 1 and 2; Figs. 2 and 3). Plasma KS concentrations measured by ELISA increased until age 10 years, at which point they fell gradually until normalizing around age 20 years. Plasma KS measurements by LC/MS/MS were highest in the first 5 years of life, and declined gradually until reaching a plateau near 20 years of age. Urine KS concentrations were highest during the first 5 years of life. After 5 years of age, urine KS levels declined until stabilizing around 20 years of age. This age-dependent pattern was observed with both ELISA and LC/MS/MS urine KS measurements.

Bottom Line: KS is synthesized mainly in cartilage and released into circulation, making it a critical biomarker for MPS IVA to evaluate clinical course and effectiveness of therapies.No correlation was found between plasma KS measurements in controls.A moderate correlation between blood and urine KS measurements in the same individual was observed.

View Article: PubMed Central - PubMed

Affiliation: Department of Pediatrics, School of Medicine, Saint Louis University, St. Louis, MO, USA.

ABSTRACT

Background and aim: Mucopolysaccharidosis IVA (MPS IVA) leads to skeletal dysplasia through excessive storage of chondroitin-6-sulfate and keratan sulfate (KS). KS is synthesized mainly in cartilage and released into circulation, making it a critical biomarker for MPS IVA to evaluate clinical course and effectiveness of therapies. Therefore, an accurate and sensitive method is required to measure KS levels.

Material and methods: Using sandwich ELISA and liquid chromatography tandem mass spectrometry (LC/MS/MS) assays, we measured KS levels in blood and urine from MPS IVA patients and healthy controls to evaluate comparability of results. Blood (patients, n = 110; controls, n = 364) and urine (patients, n = 103; controls, n = 326) specimens were obtained.

Results: Plasma and urine KS measurements in patients were age-dependent and higher than age-matched controls. We observed a moderate correlation (r = 0.666; P < 0.001) between urine KS measurements and a weak correlation (r = 0.333; P = 0.002) between plasma KS measurements by ELISA and LC/MS/MS methods in patients. No correlation was found between plasma KS measurements in controls. The difference between KS measurements assayed by LC/MS/MS and ELISA was greater in controls than in patients. A moderate correlation between blood and urine KS measurements in the same individual was observed.

Conclusion: These findings indicate that both methods to measure blood and urine KS are suitable for diagnosis, monitoring therapies, and longitudinal assessment of the disease course in MPS IVA, but the LC/MS/MS method measures over 10 times more KS present in body fluids.

No MeSH data available.


Related in: MedlinePlus