Analytical and Bioanalytical Chemistry

, Volume 410, Issue 6, pp 1679–1688 | Cite as

An immunocapture-LC-MS-based assay for serum SPINK1 allows simultaneous quantification and detection of SPINK1 variants

  • Suvi Ravela
  • Leena Valmu
  • Mykola Domanskyy
  • Hannu Koistinen
  • Leena Kylänpää
  • Outi Lindström
  • Jakob Stenman
  • Esa Hämäläinen
  • Ulf-Håkan Stenman
  • Outi Itkonen
Research Paper


Pancreatic secretory trypsin inhibitor Kazal type 1 (SPINK1) is a 6420 Da peptide produced by the pancreas, but also by several other tissues and many tumors. Some mutations of the SPINK1 gene, like the one causing amino acid change N34S, have been shown to confer susceptibility to recurrent or chronic pancreatitis. Detection of such variants are therefore of clinical utility. So far SPINK1 variants have been determined by DNA techniques. We have developed and validated an immunocapture-liquid chromatography-mass spectrometric (IC-LC-MS) assay for the detection and quantification of serum SPINK1, N34S-SPINK1, and P55S-SPINK1. We compared this method with a time-resolved immunofluorometric assay (TR-IFMA) for serum samples and primer extension analysis of DNA samples. We used serum and DNA samples from patients with acute pancreatitis, renal cell carcinoma, or benign urological conditions. With the help of a zygosity score calculated from the respective peak areas using the formula wild-type (wt) SPINK1/(variant SPINK1 + wt SPINK1), we were able to correctly characterize the heterozygotes and homozygotes from the samples with DNA information. The score was then used to characterize the apparent zygosity of the samples with no DNA characterization. The IC-LC-MS method for SPINK1 was linear over the concentration range 0.5–1000 μg/L. The limit of quantitation (LOQ) was 0.5 μg/L. The IC-LC-MS and the TR-IFMA assays showed good correlation. The median zygosity score was 1.00 (95% CI 0.98–1.01, n = 11), 0.55 (95% CI 0.43–0.61, n = 14), and 0.05 (range 0.04–0.07, n = 3) for individuals found to be wt, heterozygous, and homozygous, respectively, for the N34S-SPINK1 variant by DNA analysis. When DNA samples are not available, this assay facilitates identification of the N34S- and P55S-SPINK1 variants also in archival serum samples.


Immunocapture TATI SPINK1 Quantitative LC-MS assay 



The authors thank Ms. Maarit Leinimaa and Ms. Marianne Niemelä for expert technical assistance. This work was supported by the grants from the Academy of Finland, Finska läkaresällskapet, Sigrid Jusélius Foundation, Medical Faculty of the University of Helsinki, Biomedicum Helsinki Foundation, and Timo Lehtonen Urology Fund.

Compliance with ethical standards

S. Ravela and L. Valmu work currently at ThermoFisher Scientific, but S. Ravela was a University of Helsinki employee and L. Valmu was a Finnish Red Cross employee at the time of the study. ThermoFisher Scientific has not participated nor supported this study. The authors have no other conflict of interests.

The study was approved by the ethical committee of Helsinki University Central Hospital, Finland. Informed consent was obtained from all individuals.


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Copyright information

© Springer-Verlag GmbH Germany, part of Springer Nature 2018

Authors and Affiliations

  • Suvi Ravela
    • 1
    • 2
  • Leena Valmu
    • 1
    • 2
  • Mykola Domanskyy
    • 1
  • Hannu Koistinen
    • 1
  • Leena Kylänpää
    • 3
  • Outi Lindström
    • 3
  • Jakob Stenman
    • 4
    • 5
  • Esa Hämäläinen
    • 6
  • Ulf-Håkan Stenman
    • 1
    • 6
  • Outi Itkonen
    • 1
    • 6
  1. 1.Department of Clinical ChemistryUniversity of Helsinki and Helsinki University Central HospitalHelsinkiFinland
  2. 2.Thermo Fisher ScientificVantaaFinland
  3. 3.Department of SurgeryHelsinki University Central HospitalHelsinkiFinland
  4. 4.Institute for Molecular Medicine FinlandHelsinkiFinland
  5. 5.Department of Women’s and Children’s HealthKarolinska InstitutetStockholmSweden
  6. 6.HUSLABHelsinki University Central HospitalHelsinkiFinland

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