Electrophoresis double label

Apart from the wide ran of staining techniques, fluore nt and radioactive labels are often used. FITC is used for protein detection following SDS-electrophoresis, e.g. Rat hemoglobin fractions have been assayed by labeling with Double labeling with and makes it possible to detect proteins from two different... 

Fig. 1. Altered products of mitochondrial protein synthesis in strain 73/1. Incorporation of labeled leucine in the presence of CHX by whole cells mitochondria were isolated, proteins solubilized with DOC, dissociated and analyzed by electrophoresis in 10% acrylamide gels in the presence of SDS data are cpm found in 1-mm slices. (A) CAP-sensitive incorporation by wild-type [CHX alone - (CHX -l- CAP)]. (B) Ditto by 73/1. (C) Double-labeling experiment wild-type labeled with [ Cjleucine, 73/1 with PHjleucine, both in presence of CHX, mixed and analyzed as described.

In order to define the translation site of the individual cytochrome oxidase subunits, cytochrome oxidase was purified from double-labeled cells. These had first been labeled with [ C]leucine in the absence of inhibitors, followed by labeling with [ H]leucine, either in the absence of inhibitors or in the presence of cycloheximide or chloramphenicol (see Section 2). The purified cytochrome oxidase was subsequently separated into subunits by gel electrophoresis in dodecylsulfate. The distribution of and H radioactivity was analyzed (Table I, Fig. 7). 

Fig. 4. Double-label gel profiles of isolated mitochondria obtained during petite induction. Log-phase cultures of 55-R5-3C and 1121 were grown on 2% galactose for 5-6 doublings at 18°C (A), or at 28°C in the presence of 3 mg per ml of chloramphenicol (B), and were labeled with [ H]leucine or [ C]leucine in the presence of 200 Mg per ml cycloheximide. After labeling, mitochondria were isolated and the protein was separated on 10% SDS polyacrylamide gels. After electrophoresis, the gels were sliced and counted. The data are normalized to the total radioactivity recovered from the gels after correction for spillover of the two isotopes. (From Weislogel and Butow. )...

Fig. 8. Gel electrophoresis of an envelope protein fraction double labeled with pHJarginine and [ CJhistidihe. The envelope fraction was treated with T4 phage lysozyme before electrophoresis.

Starting with the first IPCR study, gel electrophoresis retains its potential as a fast and easy method for end-point determination of DNA amplificate for IPCR assays [10, 24, 25, 29, 31, 35, 36, 38, 39, 64], Readout is performed by intercalation fluorescence markers (e.g., ethidium bromide) and photometric/densitometric quantification of band signal intensities. The direct addition of a double-strand specific intercalation marker to the PCR amplificate and subsequent measurement of fluorescence in microwells proved to be of insufficient sensitivity for the quantification of IPCR amplificate [37]. Alternative approaches, such as radioactive labeling during PCR and subsequent imaging [33], were carried out but are not well suited for routine clinical application because of additional methodological requirements. An advantage of gel electrophoresis is the possibility of simultaneous amplificate detection for multiplex IPCR [41] and the ability to detect nonspecific amplification products. 

Biochemically, apoptosis is characterized by the internucleosomal degradation of chromosomal DNA to form a series of double-stranded fragments that are multiples of 180 200 base pairs in length. These fragments give a characteristic DNA ladder pattern on gel electrophoresis [91, 92] and can be detected by several cytochemical methods, the most extensively used being the terminal deoxynucleotidyl transferase (TdT)-mediated biotinylated dUTP nick end labeling (TUNEL) [93-95], The detection of ladder pattern and TUNEL positivity has been adopted as a marker of apoptosis. 

Kolkman, A., Dieksen, E. H., Slijpee, M., Heck, A. J. (2005). Double standards in quantitative proteomics direct comparative assessment of difference in gel electrophoresis and metabolic stable isotope labeling. Mol. Cell Proteomics 4, 255-266. 

Heterogeneous immunoassay in which a piece of single- or double-stranded DNA is used as a label for an antibody in a sandwich assay. Bound DNA label is amplified using the polymerase chain reaction (PCR). The amplified DNA product is separated by gel electrophoresis and quantitated by densitometric scanning of an ethidium stained gel. 

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