By Pauline M. Doran
This quantity goals to explain essentially and intimately the main sensible abilities occupied with cartilage tissue engineering. equipment are defined for isolation and growth of chondrocytes and stem cells; differentiation; synthesis and alertness of third-dimensional scaffolds; layout and operation of bioreactors; in vivo checking out of engineered constructs; and molecular and sensible research of cartilage cells and tissues. usually used applied sciences are lined, in addition to more moderen advances in inspirational parts resembling ‘smart’ biomaterial improvement, novel bioreactor layout, -omics research, and genetic manipulation of matrix synthesis. the main approaches mentioned both underpin the development already completed in cartilage tissue engineering or are indicative of the course of destiny study within the quarter. Written within the hugely winning Methods in Molecular Biology series structure, chapters comprise introductions to their respective themes, lists of the mandatory fabrics and reagents, with ease reproducible step by step laboratory protocols, and tips about troubleshooting and heading off identified pitfalls.
Authoritative and state of the art, Cartilage Tissue Engineering: equipment and Protocols is an invaluable source that informs the medical group concerning the experimental paintings masking a vast variety of pursuits for cartilage synthesis and regeneration.
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Extra info for Cartilage Tissue Engineering: Methods and Protocols
32 Kifah Shahin et al. 6. Transfer the minced tissue to 50-mL centrifuge tubes containing digestion solution (300 mg of tissue per 10 mL of solution per tube) using scalpels. Using a 5-mL pipette, rinse the Petri dish with 1–2 mL of digestion solution and aspirate to transfer the remaining tissue pieces (see Note 10). Place the centrifuge tubes horizontally on their sides in a CO2 incubator, making sure that all tissue pieces are in the solution. Incubate at 37 °C for 14–16 h. Filter the digest through a sterile 150-μm nylon sieve to remove tissue debris and collect the released cells.
Schrobback K, Klein TJ, Schuetz M et al (2011) Adult human articular chondrocytes in a microcarrier-based culture system: expansion and redifferentiation. J Orthop Res 29:539–546 8. Jakob M, Démarteau O, Schäfer D et al (2003) Enzymatic digestion of adult human articular cartilage yields a small fraction of the total available cells. Dehne T, Karlsson C, Ringe J et al (2009) Chondrogenic differentiation potential of osteoarthritic chondrocytes and their possible use in matrix-associated autologous chondrocyte transplantation.
2. Add 50 μl of methylene blue solution and incubate for 10 min at room temperature. 3. Count the stained cells using a hemocytometer. Only count the regular, evenly stained cells. The cells are now considered to be at passage 0. 4 Counting Mononuclear Cells: Method 2 47 Alternatively, the number of isolated mononuclear cells may be determined using a Sceptre cell counter (Merck Millipore). To count with the Sceptre system: 1. Dilute the cell suspension 1:20 in PBS (50 μl of cell suspension: 950 μl of PBS).
Cartilage Tissue Engineering: Methods and Protocols by Pauline M. Doran