A reporter-cell assay for the detection of BMP-2 immobilized on porous and nonporous materials

M. Wiemann, H. P. Jennissen, H. Rumpf, L. Winkler, M. Chatzinikolaidou, I. Schmitz, D. Bingmann

Research output: Contribution to journalArticlepeer-review

22 Citations (Scopus)

Abstract

Human recombinant bone morphogenetic protein-2 (rhBMP-2) immobilized on the surface of metal implants can facilitate osseointegration. Here, we describe a cell reporter assay useful for quantifying small amounts of immobilized rhBMP-2 on various materials. The peptide was dotted and heat-fixed on titanium, 316L stainless steel, nitrocellulose, or glass, and its distribution was monitored by in situ biotinylation followed by detection with the avidin-biotin method. Bioactivity of rhBMP-2 was demonstrated by means of a confluent layer of osteoblastic MC3T3-E1 cells that evenly covered rhBMP-2-free and rhBMP-2-loaded surface areas, as shown with epifluorescence microscopy of calcein acetoxymethyl (AM)-loaded cells. Expression of osteocalcin, fibronectin, actin, and vimentin increased where cells were located on rhBMP-2 dotted areas, but the signal:noise ratio was too low to bioassay the peptide. However, local pronounced expression of alkaline phosphatase was used to quantify BMP-2 in the range of 5-80 ng/dot by means of a cytochemical color reaction for alkaline phosphatase and image analysis of resulting dots. The lower detection limit was in the order nitrocellulose > glass > titanium > 316L steel. We conclude that the cell reporter assay is useful to assess biological activity of rhBMP-2 even after immobilization on three-dimensional implant materials.

Original languageEnglish
Pages (from-to)119-127
Number of pages9
JournalJournal of Biomedical Materials Research
Volume62
Issue number1
DOIs
Publication statusPublished - 2002
Externally publishedYes

Keywords

  • Bioassay
  • Bone implant
  • MC3T3-E1 cells
  • Osteoblast
  • Surface coating

ASJC Scopus subject areas

  • Biomaterials
  • Biomedical Engineering

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