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Discrimination of grade 2 and 3 cervical intraepithelial neoplasia by means of analysis of water soluble proteins recovered from cervical biopsies

Kai-Erik Uleberg134, Ane Cecilie Munk12, Cato Brede6, Einar Gudlaugsson14, Bianca van Diermen1, Ivar Skaland1, Anais Malpica5, Emiel AM Janssen1, Anne Hjelle4 and Jan PA Baak13*

Author Affiliations

1 Pathology Department, Stavanger University Hospital, Armauer Hansen Road 20, Stavanger, Norway

2 Gynaecology Department, Stavanger University Hospital, Armauer Hansen Road 20, Stavanger, Norway

3 The Gade Institute, University of Bergen, Bergen, Norway

4 International Research Institute of Stavanger (IRIS), Stavanger, Norway

5 Departments of Pathology and Gynecologic Oncology, The University of Texas M. D. Anderson Cancer Center, Houston, TX

6 Department of Medical Biochemistry, Stavanger University Hospital, Armauer Hansen Road 20, Stavanger, Norway

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Proteome Science 2011, 9:36  doi:10.1186/1477-5956-9-36

Published: 28 June 2011



Cervical intraepithelial neoplasia (CIN) grades 2 and 3 are usually grouped and treated in the same way as "high grade", in spite of their different risk to cancer progression and spontaneous regression rates. CIN2-3 is usually diagnosed in formaldehyde-fixed paraffin embedded (FFPE) punch biopsies. This procedure virtually eliminates the availability of water-soluble proteins which could have diagnostic and prognostic value.


To investigate whether a water-soluble protein-saving biopsy processing method followed by a proteomic analysis of supernatant samples using LC-MS/MS (LTQ Orbitrap) can be used to distinguish between CIN2 and CIN3.


Fresh cervical punch biopsies from 20 women were incubated in RPMI1640 medium for 24 hours at 4°C for protein extraction and subsequently subjected to standard FFPE processing. P16 and Ki67-supported histologic consensus review CIN grade (CIN2, n = 10, CIN3, n = 10) was assessed by independent gynecological pathologists. The biopsy supernatants were depleted of 7 high abundance proteins prior to uni-dimensional LC-MS/MS analysis for protein identifications.


The age of the patients ranged from 25-40 years (median 29.7), and mean protein concentration was 0.81 mg/ml (range 0.55 - 1.14). After application of multistep identification criteria, 114 proteins were identified, including proteins like vimentin, actin, transthyretin, apolipoprotein A-1, Heat Shock protein beta 1, vitamin D binding protein and different cytokeratins. The identified proteins are annotated to metabolic processes (36%), signal transduction (27%), cell cycle processes (15%) and trafficking/transport (9%). Using binary logistic regression, Cytokeratin 2 was found to have the strongest independent discriminatory power resulting in 90% overall correct classification.


114 proteins were identified in supernatants from fresh cervical biopsies and many differed between CIN2 and 3. Cytokeratin 2 is the strongest discriminator with 90% overall correct classifications.

cervical intraepithelial neoplasia; CIN; proteomics; LTQ-Orbitrap; mass spectrometry