SummaryObjective: The need for an alternative to fetal bovine serum (FBS) is known to scientists and users involved in cell therapy or advanced therapy medicinal products. Human serum (huS) and platelet lysate (hPL) can be used as alternatives resulting in similar or even superior results concerning cell expansion. Methods: We developed protocols for the production of huS and two types of hPL and tested them in the expansion of human fibroblasts and adipose tissue-derived stem cells (ASC). Quality control included cell counts (platelets, red and white blood cells), sterility testing, pH levels, total protein concentrations and growth factor levels. ASC and fibroblasts were expanded for three passages in media supplemented with FBS, huS or hPL and evaluated microscopically. Proliferation in terms of population doubling times (PDT) was determined. In case of ASC, differentiation was performed as well. Results: All three alternatives demonstrated shorter PDT for fibroblasts and ASC compared to FBS. Furthermore, ASC maintained their differentiation potential. Conclusion: We conclude that hPL and huS can be used as alternatives to FBS for the cultivation and expansion of cells intended for human use.

1.
www.clinicaltrials.gov/ct2/results?term = cell+therapy.
2.
Martin MJ, Muotri A, Gage F, Varki A: Human embryonic stem cells express an immunogenic non-human sialic acid. Nat Med 2005;11:228-232.
3.
Tekkatte C, Gunasingh GP, Cherian KM, Sankaranarayanan K: ‘Humanized' stem cell culture techniques: the animal serum controversy. Stem Cells Int 2011;2011:504723.
4.
Note for Guidance on minimizing the risk of transmitting animal spongiform encephalopathy agents via human and veterinary medicinal products (EMA/410/01 rev.3). Official Journal of the European Union 2011/C 73/01 www.emea.europa.eu/docs/en_GB/document_library/Scientific_guide-line/2009/09/WC500003675.pdf.
5.
Bieback K, Hecker A, Kocaömer A, Lannert H, Schallmoser K, Strunk D, Klüter H: Human alternatives to fetal bovine serum for the expansion of mesenchymal stromal cells from bone marrow. Stem Cells 2009;27:2331-2341.
6.
Pratama G, Vaghjiani V, Tee JY, Liu YH, Chan J, Tan C, Murthi P, Gargett C, Manuelpillai U: Changes in culture expanded human amniotic epithelial cells: implications for potential therapeutic applications. PLoS One 2011;6:e26136.
7.
Liu Y, Kalén A, Risto O, Wahlström O: Fibroblast proliferation due to exposure to a platelet concentrate in vitro is pH dependent. Wound Rep Reg 2002;10:336-340.
8.
Rauch C, Feifel E, Amann EM, Spötl HP, Schennach H, Pfaller W, Gstraunthaler G: Alternatives to the use of fetal bovine serum: human platelet lysates as a serum substitute in cell culture media. ALTEX 2011;28:305-316.
9.
Blande IS, Bassaneze V, Lavini-Ramos C, Fae KC, Kalil J, Miyakawa AA, Schettert IT, Krieger JE: Adipose tissue mesenchymal stem cell expansion in animal serum-free medium supplemented with autologous human platelet lysate. Transfusion 2009;49:2680-2685.
10.
Chieregato K, Castegnaro S, Madeo D, Astori G, Pegoraro M, Rodeghiero F: Epidermal growth factor, basic fibroblast growth factor and platelet-derived growth factor-bb can substitute for fetal bovine serum and compete with human platelet-rich plasma in the ex-vivo expansion of mesenchymal stromal cells derived from adipose tissue. Cytotherapy 2011;13:933-943.
11.
Burnouf T, Tseng YH, Kuo YP, Su CY: Solvent/detergent treatment of platelet concentrates enhances the release of growth factors. Transfusion 2008;48:1090-1098.
12.
Hildner F, Eder MJ, Hofer K, Aberl J, Redl H, van Griensven M, Gabriel C, Peterbauer-Scherb A: Human platelet lysate successfully promotes proliferation and subsequent chondrogenic differentiation of adipose-derived stem cells: a comparison with articular chondrocytes. J Tissue Eng Regen Med 2013; doi: 10.1002/term.1649.
13.
Wolbank S, Peterbauer A, Wassermann E, Hennerbichler S, Voglauer R, van Griensven M, Duba HC, Gabriel C, Redl: Labelling of human adipose-derived stem cells for non-invasive in vivo cell tracking. Cell Tissue Bank 2007;8:163-177.
14.
Pittenger MF, AM Mackay AM, Beck SC, Jaiswal RK, Douglas R, Mosca JD, Moorman MA, Simonetti DW, Craig S, Marshak DR: Multilineage potential of adult human mesenchymal stem cells. Science 1999;284:143-147.
15.
de Girolamo L, Sartori MF, Albisetti W, Brini AT: Osteogenic differentiation of human adipose-derived stem cells: comparison of two different inductive media. J Tissue Eng Regen Med 2007;1:154-157.
16.
Wolbank S, Peterbauer A, Fahrner M, Hennerbichler S, van Griensven M, Stadler G, Redl H, Gabriel C: Dose-dependent immunomodulatory effect of human stem cells from amniotic membrane: a comparison with human mesenchymal stem cells from adipose tissue. Tissue Eng 2007;13:1173-1183.
17.
Wolbank S, Stadler G, Peterbauer A, Gillich A, Karbiener M, Streubel B, Wieser M, Katinger H, van Griensven M, Redl H, Gabriel C, Grillari J, Grillari-Voglauer R: Telomerase immortalized human amnion- and adipose-derived mesenchymal stem cells: maintenance of differentiation and immunomodulatory characteristics. Tissue Eng Part A 2009;15:1843-1854.
18.
Plöderl K, Strasser C, Hennerbichler S, Peterbauer-Scherb A, Gabriel C: Development and validation of a production process of platelet lysate for autologous use. Platelets 2011;22:204-209.
19.
Amable PR, Carias RB, Teixeira MV, da Cruz Pacheco I, Corrêa do Amaral RJ, Granjeiro JM, Borojevic R: Platelet-rich plasma preparation for regenerative medicine: optimization and quantification of cytokines and growth factors. Stem Cell Res Ther 2013;4:67.
20.
Schallmoser K, Rohde E, Reinisch A, Bartmann C, Thaler D, Drexler C, Obenauf AC, Lanzer G, Linkesch W, Strunk D: Rapid large-scale expansion of functional mesenchymal stem cells from unmanipulated bone marrow without animal serum. Tissue Eng Part C Methods 2008;14:185-196.
21.
Lohmann M, Walenda G, Hemeda H, Joussen S, Drescher W, Jockenhoevel S, Hutschenreuter G, Zenke M, Wagner W: Donor age of human platelet lysate affects proliferation and differentiation of mesenchymal stem cells. PLoS ONE 2012;7:e37839.
22.
Zaragosi LE, Ailhaud G, Dani C: Autocrine fibroblast growth factor 2 signaling is critical for self-renewal of human multipotent adipose-derived stem cells. Stem Cells 2006;24:2412-2419.
23.
Kemptner J, Witzeneder K, Hennerbichler S, Gabriel C: Proteomic analysis of human derived cell culture supplements. Transfus Med Hemother 2013;40(6):413-415.
24.
Josh F, Kobe K, Tobita M, Tanaka R, Suzuki K, Ono K, Hyakusoku H, Mizuno H: Accelerated and safe proliferation of human adipose-derived stem cells in medium supplemented with human serum. J Nippon Med Sch 2012;79:444-452.
25.
Kocaoemer A, Kern S, Kluter H, Bieback K: Human AB serum and thrombin-activated platelet-rich plasma are suitable alternatives to fetal calf serum for the expansion of mesenchymal stem cells from adipose tissue. Stem Cells 2007;25:1270-1278.
26.
Lai WT, Krishnappa V, Phinney DG: Fibroblast growth factor 2 (Fgf2) inhibits differentiation of mesenchymal stem cells by inducing Twist2 and Spry4, blocking extracellular regulated kinase activation, and altering Fgf receptor expression levels. Stem Cells 2011;29:1102-1111.
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