Two-photon laser scanning microscopy on native cartilage and collagen membranes for tissue engineering

Jörg Martini; Katja Tönsing; Michael Dickob; Ronald Schade; Klaus Liefeith; Dario Anselmetti

doi:10.1117/12.641430

23 February 2006 Two-photon laser scanning microscopy on native cartilage and collagen membranes for tissue engineering

Jörg Martini, Katja Tönsing, Michael Dickob, Ronald Schade, Klaus Liefeith, Dario Anselmetti

Proceedings Volume 6089, Multiphoton Microscopy in the Biomedical Sciences VI; 60891N (2006) https://doi.org/10.1117/12.641430
Event: SPIE BiOS, 2006, San Jose, California, United States

Abstract

In our experiments 2-Photon laser scanning microscopy (2PLSM) has been used to acquire 3-dimensional structural information on native unstained biological samples for tissue engineering purposes. Using near infrared (NIR) femtosecond laser pulses for 2-photon excitation and second harmonic generation (SHG) it was possible to achieve microscopic images at great depths in strongly (light) scattering collagen membranes (depth up to 300 μm) and cartilage samples (depth up to 460 μm). With the objective of optimizing the process of chondrocyte growth on collagen scaffolding materials for implantation into human knee joints, two types of samples have been investigated. (1) Both arthritic and non-arthritic bovine and human cartilage samples were examined in order to differentiate between these states and to estimate the density of chondrocytes. In particular, imaging depth, fluorescence intensity and surface topology appear promising as key information for discriminating between the non-arthritic and arthritic states. Human chondrocyte densities between 2-10⁶/cm³ and 20-10⁶/cm³, depending on the relative position of the sample under investigation within the cartilage, were measured using an automated procedure. (2) Chondrocytes which had been sown out on different types of I/III-collagen membranes, were discriminated from the scaffolding membranes on the basis of their native fluorescence emission spectra. With respect to the different membranes, either SHG signals from the collagen fibers of the membranes or differences in the emission spectra of the chondrocytes and the scaffolding collagenes were used to identify chondrocytes and membranes.

Citation Download Citation

Jörg Martini, Katja Tönsing, Michael Dickob, Ronald Schade, Klaus Liefeith, and Dario Anselmetti "Two-photon laser scanning microscopy on native cartilage and collagen membranes for tissue engineering", Proc. SPIE 6089, Multiphoton Microscopy in the Biomedical Sciences VI, 60891N (23 February 2006); https://doi.org/10.1117/12.641430

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