Adhesion aggregation assay

The methods described below outline three dynamic adhesion/aggregation assays used to assess the in vitro and/or ex vivo efficacy of platelet antagonists (1) a viscometric-flow cytometric assay to measure shear-induced platelet-platelet aggregation in the bulk phase, (2) a perfusion chamber coupled with a computerized videomicroscopy system to visualize in real time and quantify (a) the adhesion and subsequent aggregation of platelets flowing over an immobilized substrate (e.g. extracellular matrix protein) and (b) free-flowing monocytic cell adhesion to immobilized platelets. 

The NF2 gene is a tumor suppressor on chromosome 22. Loss of expression of the NF2 protein product, Merlin (Schwannomin), is associated with both sporadic and NF2-related schwannomas.Merlin is a cytoskeleton-associated tumor suppressor protein regulated by phosphorylation at serine 518 (S518). Unphosphorylated Merlin restricts cell proliferation by inhibiting Rac and p21-activated kinase (Pak). In Merlin-deficient schwannoma cells, Rac causes nonfunctional intercellular adhesion in aggregation assays that could cause increased proliferation rates owing to loss of contact inhibition. 1 ... 

The brain CS-PGs neurocan and phosphacan were recently shown to interact with cell adhesion molecules. The localization of neurocan- and phosphacan-immunoreactivities overlap extensively with that of tenascin, Ng-CAM and N-CAM in embryonic rat spinal cord and early postnatal rat cerebellum, respectively (Grumet et al., 1994 Friedlander et al., 1994 Milev et al., 1994). Neurocan and phosphacan interact with N-CAM, Ng-CAM and tenascin as shown by aggregation assays... 

Platelets are then washed via centrifugation (1,100 x g for 10 min) and resuspended at 2 x 10s/mL in HEPES-Tyrode buffer, and kept at room temperature for no longer than 4 hours before use in aggregation/adhesion assays. 

Aggregation of cells in suspension. The method was originally set up by Orr and Roseman (1969), and next modified by Tao et al. (1983) to measure differences in homotypic adhesiveness due to cell surface carbohydrates changes. A similar assay was also set up by others in order to evaluate how different parameters could influence the aggregation ability of BHK cells in culture (Urushihara et al., 1976 Takeichi, 1977). These methods measure the propensity of a... 

Interactions between tumor cells and platelets can be analyzed by different methods. The ability of tumor cells to induce platelet aggregation ( tumor cells induced platelet aggregation TCIPA) is evaluated by the turbidometric assay (Menter et al., 1987 Watanabe et al., 1988 Sugimoto et al., 1991 Tang et al., 1993 Belloc et al., 1995). Besides, the ability of tumor cells to bind platelets can be evaluated by the classic adhesion assay (see below). 

In vitro platelet-tumor cell adhesion assay If the ability of tumor cells to induce platelet aggregation has a prominent role in metastasis formation, it could be expected that its inhibition by specific drugs would reduce the amount of metastases produced by tumor cells with a pronounced TCIPA activity. 

The assay was originally described by Nicolson and Winkelhake (1975) in order to test the specific adhesion ability of B16 murine melanoma cells selected in vivo for lung-specific colonization (Fidler, 1973a). The test measures the ability of a tumor-cell suspension to bind to and aggregate a similar suspension of organ-derived host cells. The obtained results showed that such heterotypic aggregation occurred in a specific way, and it was related to the preferred organ site for implantation. 

Muggli R, Baumgartner HR, Tsdiopp TB, Keller H Automated microdensitometry and protein assays as a measure for platelet adhesion and aggregation on collagen-coated slides under controlled flow conditions. J Ld> Clin Med 95 195-207, 1980. 

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