{"alternative_title":["Neuromethods"],"title":"High-Resolution localization and quantitation of membrane proteins by SDS-digested freeze-fracture replica labeling (SDS-FRL)","quality_controlled":"1","status":"public","related_material":{"record":[{"status":"public","relation":"dissertation_contains","id":"9562"}]},"publisher":"Humana","oa_version":"None","date_created":"2021-07-30T09:34:56Z","user_id":"D865714E-FA4E-11E9-B85B-F5C5E5697425","series_title":"Neuromethods","department":[{"_id":"RySh"},{"_id":"EM-Fac"}],"abstract":[{"text":"High-resolution visualization and quantification of membrane proteins contribute to the understanding of their functions and the roles they play in physiological and pathological conditions. Sodium dodecyl sulfate-digested freeze-fracture replica labeling (SDS-FRL) is a powerful electron microscopy method to study quantitatively the two-dimensional distribution of transmembrane proteins and their tightly associated proteins. During treatment with SDS, intracellular organelles and proteins not anchored to the replica are dissolved, whereas integral membrane proteins captured and stabilized by carbon/platinum deposition remain on the replica. Their intra- and extracellular domains become exposed on the surface of the replica, facilitating the accessibility of antibodies and, therefore, providing higher labeling efficiency than those obtained with other immunoelectron microscopy techniques. In this chapter, we describe the protocols of SDS-FRL adapted for mammalian brain samples, and optimization of the SDS treatment to increase the labeling efficiency for quantification of Cav2.1, the alpha subunit of P/Q-type voltage-dependent calcium channels utilizing deep learning algorithms.","lang":"eng"}],"date_published":"2021-07-27T00:00:00Z","publication_status":"published","month":"07","article_processing_charge":"No","date_updated":"2024-03-25T23:30:16Z","type":"book_chapter","day":"27","project":[{"call_identifier":"H2020","name":"In situ analysis of single channel subunit composition in neurons: physiological implication in synaptic plasticity and behaviour","_id":"25CA28EA-B435-11E9-9278-68D0E5697425","grant_number":"694539"},{"call_identifier":"H2020","grant_number":"720270","name":"Human Brain Project Specific Grant Agreement 1 (HBP SGA 1)","_id":"25CBA828-B435-11E9-9278-68D0E5697425"}],"ddc":["573"],"keyword":["Freeze-fracture replica: Deep learning","Immunogold labeling","Integral membrane protein","Electron microscopy"],"acknowledgement":"This work was supported by the European Union (European Research Council Advanced grant no. 694539 and Human Brain Project Ref. 720270 to R. S.) and the Austrian Academy of Sciences (DOC fellowship to D.K.).","year":"2021","intvolume":"       169","volume":169,"_id":"9756","language":[{"iso":"eng"}],"citation":{"mla":"Kaufmann, Walter, et al. “High-Resolution Localization and Quantitation of Membrane Proteins by SDS-Digested Freeze-Fracture Replica Labeling (SDS-FRL).” <i> Receptor and Ion Channel Detection in the Brain</i>, vol. 169, Humana, 2021, pp. 267–83, doi:<a href=\"https://doi.org/10.1007/978-1-0716-1522-5_19\">10.1007/978-1-0716-1522-5_19</a>.","ieee":"W. Kaufmann, D. Kleindienst, H. Harada, and R. Shigemoto, “High-Resolution localization and quantitation of membrane proteins by SDS-digested freeze-fracture replica labeling (SDS-FRL),” in <i> Receptor and Ion Channel Detection in the Brain</i>, vol. 169, New York: Humana, 2021, pp. 267–283.","ama":"Kaufmann W, Kleindienst D, Harada H, Shigemoto R. High-Resolution localization and quantitation of membrane proteins by SDS-digested freeze-fracture replica labeling (SDS-FRL). In: <i> Receptor and Ion Channel Detection in the Brain</i>. Vol 169. Neuromethods. New York: Humana; 2021:267-283. doi:<a href=\"https://doi.org/10.1007/978-1-0716-1522-5_19\">10.1007/978-1-0716-1522-5_19</a>","ista":"Kaufmann W, Kleindienst D, Harada H, Shigemoto R. 2021.High-Resolution localization and quantitation of membrane proteins by SDS-digested freeze-fracture replica labeling (SDS-FRL). In:  Receptor and Ion Channel Detection in the Brain. Neuromethods, vol. 169, 267–283.","chicago":"Kaufmann, Walter, David Kleindienst, Harumi Harada, and Ryuichi Shigemoto. “High-Resolution Localization and Quantitation of Membrane Proteins by SDS-Digested Freeze-Fracture Replica Labeling (SDS-FRL).” In <i> Receptor and Ion Channel Detection in the Brain</i>, 169:267–83. Neuromethods. New York: Humana, 2021. <a href=\"https://doi.org/10.1007/978-1-0716-1522-5_19\">https://doi.org/10.1007/978-1-0716-1522-5_19</a>.","apa":"Kaufmann, W., Kleindienst, D., Harada, H., &#38; Shigemoto, R. (2021). High-Resolution localization and quantitation of membrane proteins by SDS-digested freeze-fracture replica labeling (SDS-FRL). In <i> Receptor and Ion Channel Detection in the Brain</i> (Vol. 169, pp. 267–283). New York: Humana. <a href=\"https://doi.org/10.1007/978-1-0716-1522-5_19\">https://doi.org/10.1007/978-1-0716-1522-5_19</a>","short":"W. Kaufmann, D. Kleindienst, H. Harada, R. Shigemoto, in:,  Receptor and Ion Channel Detection in the Brain, Humana, New York, 2021, pp. 267–283."},"ec_funded":1,"publication_identifier":{"eisbn":["9781071615225"],"isbn":["9781071615218"]},"has_accepted_license":"1","publication":" Receptor and Ion Channel Detection in the Brain","author":[{"full_name":"Kaufmann, Walter","id":"3F99E422-F248-11E8-B48F-1D18A9856A87","first_name":"Walter","last_name":"Kaufmann","orcid":"0000-0001-9735-5315"},{"first_name":"David","last_name":"Kleindienst","full_name":"Kleindienst, David","id":"42E121A4-F248-11E8-B48F-1D18A9856A87"},{"orcid":"0000-0001-7429-7896","id":"2E55CDF2-F248-11E8-B48F-1D18A9856A87","full_name":"Harada, Harumi","first_name":"Harumi","last_name":"Harada"},{"first_name":"Ryuichi","last_name":"Shigemoto","id":"499F3ABC-F248-11E8-B48F-1D18A9856A87","full_name":"Shigemoto, Ryuichi","orcid":"0000-0001-8761-9444"}],"page":"267-283","doi":"10.1007/978-1-0716-1522-5_19","place":"New York"}