TPS® ELISA (Cytokeratin 18 kit): For Research Use Only (RUO) Size: 1x96 wells Sensitivity: 6 U/L Dynamic Range: 10 - 1200 U/L Incubation Time: 2 hours 20 minutes Sample Type: Serum, Heparinized Plasma Sample Size: 50 µL Product Developed and Manufactured in Sweden by IDL Biotech AB
Intended Use TPS® ELISA (Cytokeratin 18 kit) is intended for quantitative determination of cytokeratin 18 in serum or heparinized plasma samples by enzyme immunoassay (ELISA). The TPS® ELISA (Cytokeratin 18 kit) is for research use only and not to be used in clinical, diagnostic or therapeutic procedures.
Assay Principle TPS® ELISA (Cytokeratin 18 kit) is a one step enzyme linked sandwich immunoassay (ELISA). Standards, controls and samples react during incubation simultaneously with a solid phase monoclonal catcher antibody and the HRP-conjugated monoclonal detector antibody (M3). After washing, the TMB substrate is added and after an incubation time the reaction is stopped. The absorbance at 450 nm is then measured. The developed color is directly proportional to the concentration of the cytokeratin 18 (Also known as CK18, K18, Keratin 18, KRT 18).
Pipette 50 µl standards, controls or samples per well. Leave two empty wells for background absorbance measurement (blank).
Add 100 µl TPS® ELISA HRP Conjugate per well except the blank wells. NOTE: Steps 1 and 2 should be performed sequentially without interruption.
Incubate for 2 h ± 10 min on a shaker at ~450 rpm. Correct setting of the shaker is important for correct results.
Prepare the wash solution by dissolving one wash tablet in 500 ml of deionized water.
Aspirate and wash the wells 3 times with 0.3 ml wash solution.
Add 200 µl TMB substrate per well, including the blank wells. Incubate in darkness for 20±1 min.
Add 50 µl Stop Solution per well. Agitate on a shaker for 1 min.
Read the absorbance at 450 nm within 30 min after addition of the Stop Solution
Calculate the cytokeratin 18 concentration (U/l) of the samples. Samples showing concentrations > 1200 U/l should be suitably diluted with TPS® ELISA Diluent (Standard 0 U/l) before repeated analysis.
Specificity: TPS® ELISA (Cytokeratin 18 kit) measures the M3-epitope on soluble cytokeratin 18 fragments. There is no detectable cross reactivity to cytokeratin 8 and 19.
Assay Background: All eucaryotic cells have cytoplasmic cytoskeletal structures known as intermediate filaments. The cytoskeletal network is responsible for the mechanical integrity of the cell and it is critical during cellular processes like cell division, motility and cell to cell contacts. At present more than 20 different cytokeratins have been identified, of which cytokeratin 8, 18 and 19 are the most abundant in simple epithelial cells. The cytokeratins are epithelial cell specific and the cytokeratin pattern is usually preserved during the transformation of normal cells into malignant cells.
Tumor markers have been shown to be useful in many different cancers for prediction of prognosis, monitoring therapy response and for early detection of recurrences. Many studies have been designed to identify prognostic factors for determination of suitable treatment and also when to change treatment. The prognostic information derived from many of the established biomarker measurements have failed to make a major impact on disease management. Different biomarkers have been proposed to be of additive value to established biomarkers like CA125, PSA, CA 15-3, CA 19-9 or CEA during follow-up to obtain better disease management. Published reports describe the use of the cytokeratin 18 marker TPS in many different cancers in addition to the established biomarkers to improve information about the actual activity of the cancer tumor.
References:
Stigbrand T et al. Epitope specificity of 30 monoclonal antibodies against cytokeratin antigens: The ISOBM TD5-1 Workshop. Tumor Biol 1998; 19:132-152.
van Dalen A et al. The prognostic significance of increasing marker levels in metastatic breast cancer patients with clinically complete remission, partial remission or stable disease. Int J Biol Markers 1998; 13: 10-15.
Kil PJ et al. Tissue Polypeptide Specific antigen (TPS) determinations before and during intermittent Maximal Androgen Blockade in patients with metastatic prostatic carcinoma. Eur Urol 2003; 43: 31-38.
Barak V et al. Clinical utility of cytokeratins as tumor markers. Clin Biochem 2004; 37: 529-540.
Rydlander L et al. Molecular characterization of a Tissue Polypeptide Specific-antigen epitope and its relationship to human cytokeratin 18. Eur J Biochem 1996; 241: 309-314.
van Dalen et al. Significance of serum CA125 and TPS antigen levels för determination of overall survival after three chemotherapy courses in ovarian cancer patients during long-term follow-up. Eur. J. Gynaec. Oncol. 2009; 30(6): 609-15.
