125I-labeled fasciculin 2: a new tool for quantitation of acetylcholinesterase densities at synaptic sites by EM-autoradiography.
Radio-iodinated fasciculin 2 (Fas2), a polypeptide anticholinesterase toxin from Mamba venom, was used as a new probe for localizing and quantifying acetylcholinesterase (AChE) at mouse neuromuscular junctions (NMJs) by quantitative electron microscope autoradiography. We demonstrate that 125I-Fas2 binds very specifically to the NMJs of mouse sternomastoid muscles, with very little binding to other regions in the muscles. Junctional AChE-site densities obtained from the autoradiograms were similar to those previously obtained for the same muscles using 3H-DFP. The use of 125I-Fas2 with EM-autoradiography is simpler and provides higher resolution and sensitivity, as well as considerably lower non-specific binding than previously attainable with 3H-DFP. The advantages and limitations of this procedure are discussed.
Anglister
L
L
Department of Anatomy and Cell Biology, Hebrew University--Hadassah Medical School, Jerusalem, Israel. lili@md2.huji.ac.il
Eichler
J
J
Szabo
M
M
Haesaert
B
B
Salpeter
M M
MM
D000110:Acetylcholinesterase / Q000032:analysis*; D000818:Animals; D001345:Autoradiography; D001665:Binding Sites; D004546:Elapid Venoms / Q000737:chemistry*; D017815:Elapidae; D004789:Enzyme Activation / Q000187:drug effects; D007457:Iodine Radioisotopes / Q000737:chemistry*; D008297:Male; D051379:Mice; D008854:Microscopy, Electron; D018482:Muscle, Skeletal / Q000737:chemistry / Q000187:drug effects / Q000201:enzymology; D011485:Protein Binding / Q000187:drug effects; D012680:Sensitivity and Specificity; D013569:Synapses / Q000737:chemistry* / Q000648:ultrastructure*; D014101:Torpedo; D014688:Venoms
D004546:Elapid Venoms; D007457:Iodine Radioisotopes; D014688:Venoms; C044937:fasciculin; D000110:Acetylcholinesterase