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A method for toxicological evaluation of biomaterials based on colony formation of V79 cells

  • Yoshihiko Kotoura
  • Takao Yamamuro
  • Jitsuhiko Shikata
  • Yoshiaki Kakutani
  • Toshiaki Kitsugi
  • Hiroya Tanaka
Original Articles

Summary

This report describes a method for cytotoxicity screening of biomaterials based on colony formation of V79 cells. For this test, two metals (titanium and nickel), two ceramics (alumina ceramic and tricalcium phosphate), and two types of polymeric material [high density polyethylene (HDP) and polyvinylchloride (PVC)] were used. Each metal and ceramic was cast into a disk and semidisk 49 mm in diameter and 1 to 2 mm thick. The HDP was molded into a petri dish and PVC was used as a thin film. The materials were sterilized by heating or with ethylene oxide and placed in plastic petri dishes, after which 8 ml cell suspension containing 100 cells were added to each dish. After 1 week, the colonies formed on the materials were fixed, stained, and then the number of colonies was counted. Titanium, alumina ceramic, and HDP showed no differences from the controls in terms of colonies. On the disks and the semidisks of nickel and tricalcium phosphate and on the thin disks of PVC, however, no colonies were detected. The V79 cells used in this experiment showed a rapid and logarithmically stable growth curve and such a high rate of colony formation as to form visible noticeable colonies, and were therefore suitable cells for screening test the cytotoxicity of biomaterials. Unlike other previously reported methods of in vitro cytotoxicity testing, this method permits assay of colonies formed from a single cell after proliferation directly on the materials. Moreover, the test with semidisks permits simple screening to assess the cytotoxicity is caused by either the chemical substances or the physical properties of the materials. Furthermore, since colonies of V79 cells are formed even on metals, it is generally easy to evaluate quantitatively the cytotoxicity of solid materials. As it has more advantages than other methods, this screening method appears suitable for the cytotoxicity test of biomaterials.

Keywords

Colony Formation Ethylene Oxide High Density Polyethylene Tricalcium Phosphate High Density Polyethylene 
These keywords were added by machine and not by the authors. This process is experimental and the keywords may be updated as the learning algorithm improves.

Zusammenfassung

Dieser Bericht beschreibt eine Methode zur zytotoxischen Bestimmung von Biomaterialien mittels Kolonienbildung von V79-Zellen. Für den Test wurden zwei metallische (Titan und Nickel), zwei keramische (Aluminiumoxidkeramik und Tricalciumphosphatkeramik) und zwei plastische („High density polyethylene” und „Polyvinylchroride”) Materialien verwendet. Die metallischen und keramischen Materialien wurden jeweils in Form von Scheiben und Halbscheiben von 49 mm Durchmesser und 1–2 mm Stärke gegossen. HDP wurde wie Petrischale geformt und PVC wurde mit Form von dünnen rundlichem Film getestet. Nach der Hitzesterilisation oder Gassterilisation mit Äthylenoxid wurden die Materialien in Petrischalen aus Plastik gelegt. Danach wurde in jede Schale 8 Milliliter Zellsuspension mit 100 Zellen gegeben. Nach einer Woche wurden die Kolonien, die sich auf den Materialien gebildet hatten, fixiert, gefärbt und gezählt. Titan, Aluminiumoxidkeramik und HDP wiesen bezüglich Kolonienbildung keinen Unterschied zu den Kontrollen auf. Hingegen konnten auf den Scheiben und Halbscheiben aus Nickel und Tricalciumphosphatkeramik und auf den Filmen von PVC keine Kolonien festgestellt werden. Die hier verwendeten V79-Zellen zeichnen sich durch ein rapides und stetiges logarithmisches Wachstum aus. Die Rate der Kolonienbildung ist dermaßen hoch, daß sich nach einer Woche mit bloßem Auge erkennbare Kolonien gebildet hatten. Daher eignen sich diese Zellen für den Test zur Bestimmung der Zytotoxität von Biomaterialien. Im Unterschied zu anderen berichteten Methoden erlaubt die hier beschriebene die Untersuchung von Kolonien, die sich aus einer einzigen Zelle direkt auf dem Material ausbreiten. Zudem erlaubt der Test mit den Halbscheiben einen einfachen Rückschluß auf die Ursache der Zytotoxität: sind es die chemischen Substanzen oder die physikalischen Eigenschaften des Materials. Da sich V79-

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Copyright information

© Springer-Verlag 1985

Authors and Affiliations

  • Yoshihiko Kotoura
    • 1
  • Takao Yamamuro
    • 1
  • Jitsuhiko Shikata
    • 1
  • Yoshiaki Kakutani
    • 1
  • Toshiaki Kitsugi
    • 1
  • Hiroya Tanaka
    • 1
  1. 1.Department of Orthopedic Surgery, Faculty of MedicineKyoto UniversitySakyoku, KyotoJapan

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