MYRIAM ANGÉLICA DE LA GARZA RAMOS,CASIANO DEL ANGEL MOSQUEDA
申请号：
MX2013001461
公开号：
MX358452B
申请日:
2013.02.06
申请国别(地区):
MX
年份:
2018
代理人:
摘要：
The tissue disaggregation techniques are used for separating groups of cells from the extracellular matrix for carrying out an efficient spread and expansion under suitable conditions of cellular culture. The invention refers to a hybrid method for the tissue disaggregation of the human dental pulp, using a mechanical dissociator combined with the action of at least an enzyme. The mesenchymal stem cells were obtained from the human dental pulp by hybrid disaggregation and magnetic separation of the tissue using magnetic cells CD44. Flow cytometry and confocal laser microscopy were used for analyzing the immunophenotype of the isolated cells, obtaining positive results in the identification of CD44-FITC+ and recording negative results for CD45-APC-. The hybrid method for tissue disaggregation of the present invention is useful for obtaining a main population of viable mesenchymal stem cells from the human dental pulp, which facilitates the expansion thereof under culture and maintains the undifferentiated state of said cells for the subsequent application in tissue engineering and cellular therapy.La presente invención se refiere a un método mejorado de disgregación de pulpa dental humana la mejora comprende la pulpa dental es disgregada mecánicamente en un disgregador mecánico donde se corre al menos un ciclo con una duración de entre 20 y 45 segundos, la pulpa dental obtenida es disgregada enzimáticamente adicionando cologenasa en una concentración de 1 a 2 mg/ml, y dispasa en una concentración de 2 a 3 mg/ml, ambas concentraciones considerando el volumen total dentro del tubo aspas giratorias, la pulpa dental es incubada con las enzimas en un tiempo de 30 a 60 minutos, la pulpa dental es homogenizada en un solo evento durante al menos 5 minutos, y en la etapa I) la suspensión celular es centrifugada a una velocidad de entre 250 y 350 xg durante al menos 10 minutos.
