Bone marrow harvest in dogs: model for acquisition of the total fraction of mononuclears cells/Colheita de medula ossea em caes: modelo para obtencao da fracao total de celulas mononucleares

In the present research a new protocol to harvest 10ml [kg.sup.-1] femoral bone marrow (BM) was developed to allow isolation, quantification and to test the mononuclear cell (MC) fraction viability. Forty male or female stray dogs, aging and weighting around two years old and 10kg respectively, were submitted to aseptic bone marrow harvest in a surgical environment. To achieve an ideal cell count of MC, an anatomical Steis needle was used during the procedure, which favored the indicated volume harvest in a shorter period of time without interfering cellular viability. A bone marrow collection kit was also used to filter bone fragments while maintaining harvested MC integrity during blood packaging. Meanwhile BM harvesting was conducted, animals peripheral blood collection was performed (pre, trans and post-operatory) to hematological evaluations and autologous blood transfusion was made to overcome the increased hemoglobin fall that takes place in the initial harvesting moments. The harvested and filtered BM was slowly placed over a Histopaque density gradient (1.077g [ml.sup.-1]). The material was centrifuged at 440g x for 30 minutes. The cellular ring was harvested, washed and three times centrifuged in saline 0.9%, DMEM and autologous sterile serum. Cellular ring count was conducted in neubauer chamber and its viability was performed with vital dye. In this study was possible to notice that with the harvested BM volume an average of 2.57 x 106 (± 1.56) MC [kg.sup.-1] was obtained and the cell viability was over 90% (96.72 ± 2.9%). It was concluded that the bone marrow kit and Steis needle with autologous serum cellular wash BM harvesting technique allow an ideal MC number isolation which can be administered in tissue lesions to enhance the regeneration process. Key words: bone marrow, cell transplantation, mononuclear cells, experimental model, dogs. No presente trabalho foi elaborada uma tecnica para protocolo de colheita de medula ossea (MO) (10ml. [kg.sup.-1]), do osso femoral, para isolamento, quantificacao e viabilidade da fracao total de celulas mononucleares (CM). Para tanto, 40 caes machos ou femeas, sem raca definida, com idade aproximada de dois anos, pesando em torno de 10kg, foram submetidos a procedimento asseptico em ambiente cirurgico para colheita de MO. Para a obtencao de uma quantidade suficiente de CM, durante o procedimento foi utilizada a agulha tipo Steis anatomica, que favoreceu a colheita de volume sanguineo em menor espaco de t