Subcellular localization of SV2 and other secretory vesicle components in PC12 cells by an efficient method of preembedding EM immunocytochemistry for cell cultures
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Subcellular localization of SV2 and other secretory vesicle components in PC12 cells by an efficient method of preembedding EM immunocytochemistry for cell cultures. / Tanner, V A; Ploug, Thorkil; Tao-Cheng, J H.
I: Journal of Histochemistry and Cytochemistry, Bind 44, Nr. 12, 12.1996, s. 1481-8.Publikation: Bidrag til tidsskrift › Tidsskriftartikel › Forskning › fagfællebedømt
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T1 - Subcellular localization of SV2 and other secretory vesicle components in PC12 cells by an efficient method of preembedding EM immunocytochemistry for cell cultures
AU - Tanner, V A
AU - Ploug, Thorkil
AU - Tao-Cheng, J H
PY - 1996/12
Y1 - 1996/12
N2 - We demonstrated the subcellular localization of SV2, a transmembrane protein associated with neuroendocrine secretory vesicles, in NGF-treated PC12 cells by preembedding EM immunocytochemistry (ICC), using a small gold probe followed by silver enhancement. The use of a multiwell chamber slide substantially improved the efficiency of the preembedding EM ICC procedures for cell cultures. The advantages and related caveats of this method are discussed. SV2 was distinctly localized on dusters of synaptic vesicles and large dense-cored vesicles (LDCV). The distribution of SV2 on these two types of secretory vesicles was compared quantitatively to that of another secretory vesicle-associated transmembrane protein, synaptophysin. In cultures under similar experimental conditions, the ratio of SV2 vs synaptophysin ICC staining on synaptic vesicle dusters was about 1:1, whereas it was about 9:1 on LDCV membranes. Furthermore, whereas SV2 is localized on the membranes of the LDCVs, chromogranin A, an acidic protein in secretory granules, is clearly in the core of the LDCVs. This is the first demonstration of these two antigens in such dose (approximately 20 nm) yet distinct compartments within a single organelle.
AB - We demonstrated the subcellular localization of SV2, a transmembrane protein associated with neuroendocrine secretory vesicles, in NGF-treated PC12 cells by preembedding EM immunocytochemistry (ICC), using a small gold probe followed by silver enhancement. The use of a multiwell chamber slide substantially improved the efficiency of the preembedding EM ICC procedures for cell cultures. The advantages and related caveats of this method are discussed. SV2 was distinctly localized on dusters of synaptic vesicles and large dense-cored vesicles (LDCV). The distribution of SV2 on these two types of secretory vesicles was compared quantitatively to that of another secretory vesicle-associated transmembrane protein, synaptophysin. In cultures under similar experimental conditions, the ratio of SV2 vs synaptophysin ICC staining on synaptic vesicle dusters was about 1:1, whereas it was about 9:1 on LDCV membranes. Furthermore, whereas SV2 is localized on the membranes of the LDCVs, chromogranin A, an acidic protein in secretory granules, is clearly in the core of the LDCVs. This is the first demonstration of these two antigens in such dose (approximately 20 nm) yet distinct compartments within a single organelle.
KW - Animals
KW - Cells, Cultured
KW - Immunohistochemistry
KW - Membrane Glycoproteins
KW - Microscopy, Electron
KW - Nerve Tissue Proteins
KW - PC12 Cells
KW - Rats
KW - Subcellular Fractions
KW - Synaptic Vesicles
KW - Synaptophysin
M3 - Journal article
C2 - 8985140
VL - 44
SP - 1481
EP - 1488
JO - Journal of Histochemistry and Cytochemistry
JF - Journal of Histochemistry and Cytochemistry
SN - 0022-1554
IS - 12
ER -
ID: 123666069