2 articles published in JoVE
An Improved Protocol to Purify and Directly Mono-Biotinylate Recombinant BDNF in a Tube for Cellular Trafficking Studies in Neurons Nicolás Stuardo1,2, Guillermo Moya-Alvarado1,3, Carolina Ramírez1,3, Giampietro Schiavo4, Francisca C. Bronfman1,3 1Department of Physiology, Faculty of Biological Sciences, Pontificia Universidad Católica de Chile, 2Department of Molecular and Cellular Biology, Faculty of Biological Sciences, Center for Aging and Regeneration (CARE UC), Pontificia Universidad Católica de Chile, 3Institute of Biomedical Sciences. Faculty of Medicine and Faculty of Life Sciences, Universidad Andrés Bello, 4Department of Neuromuscular Diseases, UCL Queen Square Institute of Neurology and UK Dementia Research Institute at UCL, University College London Campus Recombinant BDNF containing an Avi sequence (BDNFAvi) is produced in HEK293 cells in a cost-effective manner and is purified by affinity chromatography. BDNFavi is then directly mono-biotinylated with the enzyme BirA in a tube. BDNFavi and mono-biotinylated BDNFavi retain their biological activity when compared to commercially available BDNF.
Studying Organelle Dynamics in B Cells During Immune Synapse Formation Jorge Ibañez-Vega*1, Danitza Fuentes*1, Jonathan Lagos1,2, Jorge Cancino3, María Isabel Yuseff1 1Laboratory of Immune Cell Biology, Department of Cellular and Molecular Biology, Pontificia Universidad Católica de Chile, 2Faculty of Medicine, Pontificia Universidad Católica de Chile, 3Centro de Investigaciones en Biología Celular y Biomedicina, Facultad de Ciencia, Universidad San Sebastián Herein we describe two approaches to characterize cell polarization events in B lymphocytes during the formation of an IS. The first, involves quantification of organelle recruitment and cytoskeleton rearrangements at the synaptic membrane. The second is a biochemical approach, to characterize changes in composition of the centrosome, which undergoes polarization to the immune synapse.