202 research outputs found
Calvin United Church of Christ Confirmation Class, 1926
No full textA photograph of the 1926 confirmation class of Calvin United Church of Christ with Reverend Louis Bogar.1
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Bottom How-Mary Molnar, Rose Kupecz, Rev. Erivulka, Rev. Bdgar, Lola Toth,I.Tarczali
2nd. Row-Janka Balaza, Mary Varga, Elizabeth Jelinek,""",Elizabeth Meszaros,
Irene Olah, Viola Boros, Elizabeth Olah. 3rd. Row-Gus Gulyas, Joseph Rasi, Steve Olah.
Sandor Lakatos, Steve Doszpoly, Bela Veres, Millie Bertok, Paul Polyacsko, H.Orosz.
4th Row-Elek Csontos, A. Molnar, J. Varga, J. Brazlovitz,"""" v S """.AXos
Analysis of the expression profile and regional distribution of neurotransmitter receptors and ion channels in the central nervous system using histoblots
No full textThe histoblot method is a reliable and convenient way to compare the regional distribution and expression level of different proteins in brain samples without compromising the integrity of antibody binding sites by tissue fixation, which is required for conventional immunocytochemistry. Fixation introduces covalent modifications, crosslinking and/or denaturation of proteins. These chemical modifications often alter the antibody-binding sites and cross-linked molecules may hinder the access of antibody to epitopes. The direct transfer of native proteins from unfixed frozen tissue sections to an immobilizing matrix offers much improved accessibility of the transferred proteins for immunochemical analysis. The histoblot method has been successfully applied to analyse the regional distribution of several neurotransmitter receptors, ion channels and other proteins in the adult and developing brains. While this technique lacks cellular resolution, it provides high sensitivity and much improved consistency compare to conventional immunohistochemical techniques, which is essential for reliable quantitative comparisons of overall expression levels of proteins in different brain regions. Compare to conventional immunoblot analysis of protein extracts from dissected brain regions, histoblots provide more accurate and direct information about the anatomical localization of proteins. In this chapter we describe the histoblot protocol we have used for the identification of quantitative changes in a wide range of neurotransmitter receptors and ion channels in various brain regions
Investigation of neurotransmitter receptors in brain slices using cell surface biotinylation
No full textCell surface trafficking and endocytosis of neurotransmitter receptors are important regulatory mechanisms of neurotransmission. Biotinylation of plasma membrane proteins in brain slices allow their separation from those present in intracellular organelles. Membrane-impermeable, thiol-cleavable amine-reactive biotinylation reagents (e.g. EZ-link sulfo-NH-SS-biotin) form a stable covalent linkage with primary amino groups of surface exposed proteins. Following homogenisation of brain slices and solubilisation of membranes, biotin-labelled proteins can be isolated with avidin or streptavidin linked to agarose beads. Bound biotinylated proteins are released from avidin or streptavidin in the presence of reducing agents (e.g. glutathione or β-mercaptoethanol). Quantitative differences in the molecular composition of biotin-labelled (surface) and unlabelled (intracellular) protein fractions can be analysed using immunoblotting with target protein specific antibodies. While many variations of this procedure exist in the literature, in this chapter we describe the biotinylation protocol that we have applied for the investigation of quantitative changes in the cell surface expression and internalisation of ionotropic glutamate receptors in acute brain slices
Localization of Neurotransmitter Receptor and Ion Channel Proteins in Unfixed Brains Using In Situ Immunoblotting
No full textThe in situ immunoblotting (histoblot) method is a reliable and convenient way to compare the regional distribution and expression level of different proteins in brain samples without compromising the integrity of antibody binding sites by tissue fixation, which is required for conventional immunohistochemistry. Fixation introduces covalent modifications, crosslinking, and/or denaturation of proteins. These chemical modifications often alter the antibody binding sites and cross-linked molecules may hinder the access of antibody to epitopes. Therefore, several antibodies that are suitable for the investigation of native or denatured proteins fail to interact with their targets in fixed tissue samples used for immunohistochemical studies. The direct mechanical transfer of native proteins from unfixed frozen tissue sections to an immobilizing matrix (e.g., nitrocellulose membrane used for conventional immunoblotting) preserves the anatomical distribution patterns and structure of brain proteins. Also, the transferred proteins are readily accessible for immunochemical analysis on the surface of nitrocellulose membranes. Therefore, this method often enables the use of antibodies which do not recognize the target protein in fixed tissue samples. The histoblot method has been successfully applied to analyze the regional distribution of several neurotransmitter receptors, ion channels, and other proteins in the adult and developing brains. While this technique lacks cellular resolution, it provides high sensitivity and much improved consistency compare to conventional immunohistochemical techniques, which is essential for reliable quantitative comparisons of overall expression levels of proteins in different brain regions. Compared to conventional immunoblot analysis of protein extracts from dissected brain regions, histoblots provide more accurate and direct information about the anatomical localization and expression levels of proteins. In this updated chapter we describe the histoblot protocol we have used for the identification of quantitative changes in a wide range of neurotransmitter receptors and ion channels in various brain regions
Quantitative Analysis of Cell Surface Expressed, Intracellular, and Internalized Neurotransmitter Receptor Populations in Brain Slices Using Biotinylation
No full textCell surface trafficking and endocytosis of neurotransmitter receptors are important regulatory mechanisms of neurotransmission. Biotinylation of plasma membrane (PM) proteins in brain slices allows their separation from those present in intracellular organelles. An extension of this approach also enables the selective retrieval of proteins endocytosed from the plasma membrane into intracellular compartments. Membrane-impermeable, thiol-cleavable amine-reactive biotinylation reagents (e.g., EZ-link sulfo-NH-SS-biotin) form a stable covalent linkage with primary amino groups of surface-exposed proteins. Following homogenization of brain slices and solubilization of membranes, biotin-labeled proteins can be isolated with avidin (or streptavidin or NeutrAvidin) linked to a solid matrix. Bound biotinylated proteins are released from avidin in the presence of reducing agents (e.g., glutathione or β-mercaptoethanol). Quantitative differences in the molecular composition of biotin-labeled (surface or internalized) and unlabeled (intracellular) protein fractions can be analyzed separately using immunoblotting with target protein-specific antibodies. While many variations of this procedure exist in the literature, in this chapter we describe the biotinylation protocol that we have applied for the investigation of quantitative changes in the cell surface expression and internalization of ionotropic glutamate receptors in acute brain slices
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