Biomedical subjects

S Kirkeby

Publications and source records attributed to S Kirkeby.

At least 19 recordsLinked to original sources

PubMed2005

Analyses of Pseudomonas aeruginosa lectin binding to alpha-galactosylated glycans.

The specificity and binding capacity of the galactophilic lectin from the Gram negative bacterium Pseudomonas aeruginosa (PA-IL) was determined by solid phase measurements using galactosylated neoglycoproteins immobilized on microtiter plates. The bacterial lectin reacted with both short chain (monosaccharide) and long chain (pentasaccharide) glycoconjugates. Among the Galalpha1-XGal disaccharides, the highest affinity was observed towards the Galalpha1-3Gal structure. Raising the incubation temperature enhanced the lectin-polysaccharide agglutination, and it is suggested that binding to certain conformations of polysaccharides could vary between lectins with the same monocarbohydrate specificity and that this activity may, in part, be temperature dependent. Histochemical examination of lectin binding to different porcine tissues suggests a differential glycosylation of the carbohydrate antigens on endothelial cells in various parts of the vascular system. In the pancreas, PA-IL also adhered to the excretory ducts. These observations on PA-IL binding could be of importance both to determine infection foci in P. aeruginosa-mediated vacuities and to determine its role for pancreatic involvement in cystic fibrosis.

Animals↗

PubMed2004

The bacterial flora of alpha-Gal knockout mice express the alpha-Gal epitope comparable to wild type mice.

The human genome possesses pseudogenes for the enzyme alpha1,3 galactosyltransferase and hence, human cells and tissues do not express the Galalpha terminated trisaccharide structure Galalpha1-3Galbeta1-4GlcNAc, the so-called alpha-Gal epitope. Circulating antibodies specific for this carbohydrate epitope are, however, present in high amounts in humans. It has previously been hypothesized that the antibody production is induced by the presence of the alpha-Gal epitope in the cell walls of the enteric flora, especially Enterobacteriaceae spp. However, in mice, in which the epitope has been deleted by targeted mutation of the gal-transferase gene, alpha-Gal antibodies do not appear without prior immunization, although the mice through their growth probably have been exposed to a normal bacterial flora of e.g. Enterobacteriaceae spp. It is unknown whether there are different types of immune reactions to antigenic carbohydrate expressing bacteria and whether there are discrepancies in the enteric flora between these knockout mice and their wild type litter mates. In this study the enteric flora of alpha-Gal knockout and wild type mice was compared both in relation to the prevalence of different types of bacteria in the two groups of mice, as well as in relation to the expression of the epitope on the surface of Enterobacteriaceae spp. Our results showed that the enteric flora did not differ significantly between knockout and wild type mice and that it was comparable to the flora known to be present in the intestines of other mice. All Enterobacteriaceae spp. examined expressed the alpha-Gal epitope no matter whether they were isolated from knockout or wild type mice. It is, therefore, discussed whether it is more reasonable to assume that alpha-Gal antibodies in mammals that do not produce alpha1,3 galactosyltransferase such as in the knock mice and in humans are the result of another antigen stimulant than these common representatives of the enteric flora, that we isolated from the two types of mice. Possible candidates for a carrier in humans could be bacteria or viruses not isolated from barrier-bred mice.

Animals↗

PubMed2001

Binding of Griffonia simplicifolia 1 isolectin B4 (GS1 B4) to alpha-galactose antigens.

Glycoconjugates with terminal Galalpha1-3Galbeta1-4GlcNAc sequences (alpha-galactosyl epitopes, natural xenoreactive antigens) are present on various tissues in pigs and are recognized by human anti-alphagalactosyl (alphaGal) antibodies1. Hence xenotransplantation (pig-to-human) would trigger immune reactions involving complement activation and lead to the hyperactute rejection of the graft. Xenoreactive antigens are often studied by using the lectin Griffonia simplicifolia 1 isolectin B4 (GS1 B4), which shows high affinity to galactose. We here estimate the specificity of GS1 B4 for detecting various galactosyl epitopes by measuring lectin binding to neoglycoproteins, thyroglobulin and pig skeletal muscle. Enzyme linked lectin assays confirmed that GS1 B4 was highly specific to alpha-galactosylated neoglycoproteins while the lectin did not detect a beta-galactosylated ligand. The length of the sugar chains influenced the lectin-carbohydrate interaction. A monosaccharide linked to serum albumin showed higher lectin affinity than did neoglycoproteins with di- and tri-alpha-galactosyl epitopes. When the carbohydrate was extended, as in the xenoreactive pentasaccharide (Galalpha1-3Galbeta1-4GlcNAcbeta1-3Galbeta1-4Glc), the carbohydrate- lectin interaction was meagre. Not only the terminal, but also the subterminal sugar affected the lectin binding because the GS1 B4 affinity to Galalpha1-3Gal was much stronger than to Galalpha1-3GalNAc. In bovine and porcine thyroglobulin most alphaGal epitopes appear to be cryptic, but are unmasked by a heat denaturation. In pig skeletal muscle there was lectin reaction not only in the muscle capillaries, but also in the connective tissue and intracellularly in muscle fibres. In Western blots of isolated proteins from pig muscle at least three bands were strongly stained after incubation with lectin.

Animals↗

PubMed2001

Histochemical studies of the masseter, the temporal and small zygomaticomandibular, and the temporomandibular masticatory muscles from aged male and female humans. Fiber types and myosin isoforms.

The purpose of this study was to investigate the histology of two small masticatory muscles from females and males of more than 70 years of age. By using immuno- and enzyme histochemistry the muscles were characterized by their fiber types and myosin heavy chain pattern. The observations were compared with similar studies of the masseter and temporalis muscles. Previously the two small muscles have been described based solely upon their gross anatomy. One muscle originates from the anterior, deep surface of the temporal fascia and inserts in the temporal tendon: the temporo-mandibular muscle (TM). The other muscle originates from the upper part of the temporal surface of the frontal process of the zygomatic bone and the adjacent part of the frontal bone and inserts in the temporal tendon: the zygomaticomandibular muscle (ZM). In the masseter, TM, and ZM, most of the autopsy samples contained an abundant number of fibers containing neonatal myosin heavy chains while in the temporal muscle specimens, such fibers were sparse and scattered. Electrophoresis followed by immuno-staining of Western blots supported the histochemical findings. There was no obvious correspondence between fiber typing based upon ATPase activity and the neonatal myosin heavy chain content in the muscle fibers. Neither did the fibers show accordance in their content of adult slow and fast myosin heavy chains and in their content of neonatal myosin heavy chain.

Aged↗

PubMed2000

Aging affects different human muscles in various ways. An image analysis of the histomorphometric characteristics of fiber types in human masseter and vastus lateralis muscles from young adults and the very old.

This study is an attempt to objectively evaluate age-related changes in human muscles by use of histomorphometric methods. Aging in humans induces dramatic transformations in the skeletal muscles but little is known as to whether or not the aging processes per se may affect all muscles equally. In this study aging of two human muscles with different functions, origin and nerve supply is compared. Sections were cut from masseter and vastus lateralis muscles obtained from young adults aged 18-24 years and from the very old aged 90-102 years. Muscle fiber types were classified with the traditional myofibrillar ATPase staining. Various histomorphometric parameters of the different fiber types in human masseter and vastus lateralis muscle sections were obtained by image analyses to evaluate the age-related changes in the muscle fibers. The following variables were calculated: the number of each fiber type per photographed area; the area of each fiber and two indicators for the shape of the muscle fibers. In the aging muscles there was no relative preferential loss of a fiber type. High numbers of intermediate ATPase-stained fibers (IM fibers) were found in some old vastus muscles but were only sporadic in young vastus muscles. However, there was no change in the percentage distribution of intermediate ATPase-stained fibers when young and very old human masseter muscles were compared. Incubation of the sections with antimyosin antibodies showed that the IM fibers in old masseter and old vastus contained different myosin heavy chains. Thus ATPase activity and anti-myosin staining displayed a somewhat different pattern of fiber type distribution. The main changes in the shape and area indicated that type I fibers in the masseter became more circular while in the vastus they decreased significantly in size. The type II fibers in the vastus became very small and deviated significantly from circularity whereas the type II fibers in the masseter only exhibited a decrease in the size of the fibers. Histomorphometric measurements show that aging affects different human muscles in various ways.

Adenosine Triphosphatases↗

PubMed1999

Zonal variation in the distribution of an alpha 1-acid glycoprotein glycoform receptor in human adrenal cortex.

Using a histochemical technique with three different alpha 1-acid glycoprotein glycoform one glycoform specific receptor has been identified in human adrenal cortex. The receptor is associated to alpha 1-acid glycoprotein glycoform B and alpha 1-acid glycoprotein glycoform C. The glycoform specific receptor was located in the cytoplasm of glomerulosa and outer fasciculata cells. The intensity of the reaction product decreased in the fasciculata, and no staining was seen in inner fasciculata and reticularis. Inhibition with the simple sugars, mannose and GlcNAc confirmed a lectin-like reaction. The binding activity was dependent on the presence of calcium ions and not on thiol reagents. Thus the lectin-like receptor may belong to the C-type lectin family. Using an antibody to alpha 1-acid glycoprotein the presence of alpha 1-acid glycoprotein was observed in the same location as the glycoform specific receptor. The binding of alpha 1-acid glycoprotein glycoform B and alpha 1-acid glycoprotein glycoform C to the glycoform specific receptor is inhibited by the steroid hormones cortisone, aldosterone, estradiol and progesterone but not by testosterone. The pronounced changes in the distribution of AGP and its glycoform receptors during cell differentiation in the adrenal cortex suggest that AGP and its complementary lectins belong to the group of lectins which control differentiation and spatial position.

Adrenal Cortex↗

PubMed1999

Binding properties of the galactose-detecting lectin Pseudomonas aeruginosa agglutinin (PA-IL) to skeletal muscle fibres. Quantitative precipitation and precipitation inhibition assays.

Pseudomonas aeruginosa agglutinin (PA-IL) staining and the influence of various carbohydrates on lectin binding to muscle sections were investigated quantitatively using a scanning and integrating microspectrophotometre. A strong dose-dependent inhibition of PA-IL staining in the sections was recorded with galabiose (Galalpha1-4Gal) while lactose (Galbeta1-4Glc) had no inhibitory effect. The affinity of PA-IL to Galalpha1 carbohydrates was studied by ELISA using immobilized glycoconjugates in which Galalpha1 glycans were attached to bovine serum albumin or ceramide. PA-IL exhibited strong binding to both simple glycoconjugates having a single Galalpha moiety and to di- and trisaccharides with terminal Galalpha1 at the non-reducing end. In all cross-sectioned muscle fibres incubated with PA-IL, the staining was present as a honeycomb-shaped network through the entire cytoplasm. Further, a dense punctuate staining could be shown in most fibres. A similar staining pattern was noticed after incubation with a monoclonal antibody against ryanodine receptors and with biotinylated ryanodine suggesting that the network could represent the sarcoplasmic reticulum. Further, Western blots of a sarcoplasmic reticulum preparation showed multiple bands after incubation with PA-IL. It may therefore be proposed that glycoconjugates carrying terminal Galalpha1 show affinity for PA-IL and are located to the sarcoplasmic reticulum.

Adhesins, Bacterial↗

PubMed1998

Galalpha1-->4Gal-glycans are expressed on myofibrillar associated proteins.

There is evidence that glycans carrying terminal galactose residues are differently expressed in the sarcoplasm of different muscle fiber types. In this study monoclonal antibodies directed against P blood group antigens Pk: Galalpha1-4Galbeta1-4Glcbeta- and P1: Galalpha1-4Galbeta1-4GlcNAcbeta- were used to detect terminal alpha-galactosylated glycoconjugates on muscle proteins. Electrotransfer of proteins, extracted from human masseter and biceps muscles, to nitrocellulose after polyacrylamide gel electrophoresis (PAGE) and incubation with anti-Pk (CD77) consistently showed two bands with apparent molecular weights of 66 kDa and 64 kDa. In fresh frozen muscle sections from some humans there was endothelial reaction with anti-CD77 in capillaries, venules and veins but not in arterioles and arteries. In muscle samples from other humans there was no staining of endothelial cells. Formalin-fixed human muscle displayed a CD77 reaction with highest accumulation of reaction product at the periphery of the fibers. This may be explained by the presence of Pk glycoconjugates on intermediate filaments in muscle fibers. In preparations of cat masseter muscle proteins the antibodies against P1Pk antigens reacted with a 170 kDa and a 55 kDa band while in preparations of cat biceps brachii only a 55 kDa band was reactive. The specificities of the antibodies were investigated by fluorescence-activated cell sorter (FACS), alpha- and beta-galactosidase digestion and inhibitory sugars. This study indicates that glycans carrying Galalpha1-4Galbeta1- epitopes are expressed on myofibrillar associated proteins.

Aged↗

PubMed1998

Sirius red and acid fuchsin staining mechanisms.

The purpose of the study was to investigate the staining mechanism of acid fuchsin and Sirius red. Acid (poly-glutamic acid), neutral (poly-hydroxyproline) and basic (poly-arginine, poly-histidine, poly-lysine) poly-amino acids, collagen types I, II and III, and arginine- and lysine-containing histones were used as test substances applied to nitrocellulose membranes as dot blots. Five micrometer sections of granulation tissue on slides were tested in parallel. Some dots and sections were treated with chloramine-T before staining with acid fuchsin and Sirius red and some with 1 M NaOH after staining. The acid and neutral poly-amino acids were not stained, but the basic amino acids polylysine and poly-arginine, poly-amino acids containing these basic amino acids and the histones and the collagens exhibited intense staining. Oxidative deamination by chloramine-T abolished the staining and 1 M NaOH removed the staining except in the case of poly-arginine. Tissue sections treated in the same way showed a considerable decrease in staining after oxidative deamination with chloramine-T; in particular, the staining of the smaller fibers was abolished. The staining was totally removed by destaining with 1 M NaOH. Therefore, acid fuchsin and Sirius red are not selectively bound to collagen; they are also bound to other proteins containing basic amino acids, and staining to a large extent is influenced by electrostatic forces. The staining seems not to be selective for collagen, and one must account for this when quantitative conclusions are drawn from collagen methods using these stains.

Animals↗

PubMed1997

Localized agglutinin staining in muscle capillaries from normal and very old atrophic human muscle using winged bean (Psophocarpus tetragonolobus) lectin.

The winged bean (Psophocarpus tetragonolobus) agglutinin (total lectin) and its basic (WBA I) and acidic isoform (WBA II) were used to analyze capillaries in sections from human muscle. The microvessels were clearly labeled after incubation with the lectins in both normal muscle and in old muscles with age-related type II atrophy or muscle fiber grouping. Muscle fibers, nerves, and connective tissue remained unstained. The total lectin detected muscle capillaries from all blood group AB0 individuals. The isoform WBA I reacted only with blood vessels in blood group A and B individuals, while the blood vessels in blood group 0 individuals were demonstrated with WBA II. WBA I staining was inhibited by p-nitrophenyl alpha-galactopyranoside and N-acetylgalactosamine, whereas 2'-fucosyllactose and preincubation with an antibody against type-1 chain H abolished capillary staining with WBA II. The study demonstrates the usefulness of WBA as a marker of capillaries in human muscle.

Adult↗

PubMed1997

The complex-type oligosaccharide binding lectin Datura stramonium agglutinin detects type II A muscle fibres in the branchial biceps from man and cat.

Complex-type oligosaccharides were detected in the sarcoplasm of muscle fibres from cat and human biceps using lectins and anticarbohydrate antibodies. The lectin Datura stramonium agglutinin strongly stained type II A fibres as identified by myosin ATPase activity after alkaline and acid preincubation. In contrast, all muscle fibres showed a moderate coarse granular staining after incubation with Tetracarpidum conophorum agglutinin and Telfairia occidentalis agglutinin which recognize tri-antennary complex glycans poorly bound by D. stramonium agglutinin. Strong sarcoplasmic staining in all muscle fibres was obtained after incubation with an antibody against branched N-acetyllactosamine structure while an antibody against binary 2 --> 3 sialyllactosamine glycans failed to detect the muscle fibres. Treatment of the muscle sections with sialidase prior to incubation with D. stramonium agglutinin did not influence the lectin staining pattern. Staining of blots from electrophoretically separated muscle proteins obtained by homogenization, solubilization and centrifugation of small muscle pieces showed D. stramonium agglutinin binding to a number of bands ranging from 200 kDa to 30 kDa. No D. stramonium agglutinin positive bands were observed in blots from separated mitochondrial proteins while blots from sarcoplasmic reticulum separated by electrophoresis stained many bands in the range from 200 kDa to 30 kDa. It may be concluded that all muscle fibres in human and cat biceps hold intracellular non-sialylated complex-type oligosaccharides and further, that a specific tri-antennary complex-type glycoform is strongly expressed in type II A fibres as recognized by D. stramonium agglutinin. These results indicate a different glycosylation of certain myofibrillar-associated proteins in muscle fibre types.

Animals↗

PubMed1996

Lectin-like receptor for alpha 1-acid glycoprotein in the epithelium of the rat prostate gland and seminal vesicles.

BACKGROUND: A receptor for alpha 1-acid glycoprotein glycoforms AGP-B and AGP-C in the epithelium of the rat prostate gland and seminal vesicles is described. METHODS: The interaction between AGP-glycoforms and their receptor is a lectin-like interaction confirmed by inhibition of the binding by mannose and N-Acetyl-D-glucosamine. RESULTS: In vitro the receptor was also inhibited by the steroid hormones cortisone, aldosterone, progesterone, and estradiol, but not by testosterone. A significant regional variation in the expression of AGP-lectin receptor and in the localization of AGP was seen in rat prostate and seminal vesicles. The localization of the AGP lectin receptor is compared to the localization of glycoprotein AGP, and small differences are found. CONCLUSIONS: It is proposed the AGP receptors in the prostate and seminal vesicles belong to a group of lectins in the control of differentiation and organ formation.

Aldosterone↗

PubMed1996

A monoclonal anticarbohydrate antibody detecting superfast myosin in the masseter muscle.

Superfast-contracting muscle fibres (II M) were identified by ATPase staining and after incubation with an antiserum raised against myosin type II M and with an antibody raised against the Galalpha1-3Galbeta1-4GlcNAc structure. II M fibres were present in masseter muscles from cat, dog and Macaca fascicularis but not in limb muscles from the same animals and not in masseter muscles from rat, pig, cow or man. Electrophoresis and staining of blots from myosin preparations showed that the anticarbohydrate antibody detected myosin heavy chains from cat masseter but not myosin heavy chains from cat biceps. The alpha-galactose specific lectin Griffonia simplicifolia isolectin B4 (GS I B4) did not stain muscle fibres or myosin heavy chains. Therefore, the epitope on myosin heavy chains defined by the anticarbohydrate antibody is presumably not Galalpha1-3Galbeta1-4GlcNAc although the antibody staining was strongly inhibited after absorption by 10mM of this trisaccharide. Antibody staining of the muscle fibres was totally inhibited by adding 10mM p-nitrophenyl beta-D-glucuronide to the incubation medium. The results thus imply that an anticarbohydrate antibody distinctively detects a carbohydrate epitope specific for myosin in superfast contracting muscle fibres from jaw-closing muscles and confirm that this epitope is not present in other muscle fibre types. This appears to be the first report on differentiated glycosylation among myosin isoforms.

Animals↗

PubMed1996

The impact of fixatives on the binding of lectins to N-acetyl-glucosamine residues of human syncytiotrophoblast: a quantitative histochemical study.

We describe a quantitative histochemical method for demonstration of five N-acetyl-glucosamine binding lectins in the syncytiotrophoblast of human term placenta. The method employs biotinylated lectins and alkaline phosphatase-conjugated avidin. The alkaline phosphatase activity is detected by using 5-bromo-4-chloro-indoxyl phosphate as the substrate and nitroblue tetrazolium as the capture agent. The effect of 13 fixative solutions on specific lectin binding and nonspecific background staining was quantified by microspectrophotometry. Acid fixatives or fixatives containing mercuric chloride, e.g., Carnoy's and Zenker's fixatives, gave intense specific lectin binding and low background staining. Glutaraldehyde, carbodiimide, and ethanol resulted in low specific lectin binding and a very high background staining that was mainly due to endogenous placental alkaline phosphatase. Lectin binding to N-acetyl-galactosamine, mannose, galactose, and fucose was also significantly higher in sections from tissues fixed in an acid fixative compared with a neutral buffered fixative. Unfixed cryosections revealed a considerably lower degree of specific lectin binding compared with sections from fixed tissues. The activity of endogenous placental alkaline phosphatase was inhibited dose-dependently by mercuric chloride and decreased with L-phenylalanine concentration over the range of 7.8 x 10(-4) M to 5 x 10(-2) M, after which there was no further inhibition. Calf intestinal-type alkaline phosphatase conjugated to avidin was not inhibited by 5 x 10(-2) M L-phenylalanine. Endogenous placental biotin did not contribute significantly to background staining. Despite the high level of placental alkaline phsophatase, the intestinal-type alkaline phosphatase can be used as a marker enzyme in the sensitive ABC technique, provided that the nonspecific background is measured and substracted. Moreover, it is advisable to use an acid- and/or mercuric chloride-containing fixative and to add L-phenylalanine during incubation steps.

Acetylglucosamine↗

PubMed1996

Appearance and cellular distribution of lectin-like receptors for alpha 1-acid glycoprotein in the developing rat testis.

A histochemical avidin-biotin technique with three different alpha 1-acid glycoprotein glycoforms showed pronounced alterations in the cellular localization of two alpha 1-acid glycoprotein lectin-like receptors during cell differentiation in the developing rat testis. The binding of alpha 1-acid glycoprotein glycoforms to their receptors is inhibited by steroids. Testosterone, oestradiol and progesterone inhibited the binding of alpha 1-acid glycoprotein glycoform A to its receptor. Cortisone, aldosterone, oestradiol and progesterone inhibited the binding of alpha 1-acid glycoprotein glycoforms B and C to their receptor. A difference in the cellular content of alpha 1-acid glycoprotein glycoforms and alpha 1-acid glycoprotein receptors separates the spermatocytes and the early spermatids from the late spermatids. The difference in receptor composition implies a difference in the effect of different steroid hormones. The Leydig cells contained alpha 1-acid glycoprotein and lectin-like receptors for one of the glycoforms of alpha 1-acid glycoprotein from birth. The interaction between alpha 1-acid glycoprotein glycoforms and their receptors may modulate the actions of testosterone and other steroids in the testis.

Aldosterone↗

PubMed1994

Immunohistochemistry of the intercellular matrix components and the epithelio-mesenchymal junction of the human tooth germ.

The immunohistochemical localization of heparan sulphate, collagen type I, III and IV, laminin, tenascin, plasma- and cellular fibronectin was studied in tooth germs from human fetuses. The lamina basalis ameloblastica or membrana preformativa, which separates the pre-ameloblasts from the pre-dentin and dentin, contained heparan sulphate, collagen type IV, laminin and fibronectin. Enamel reacted with antifibronectin, but the reaction varied depending on the type of fibronectin and the source of antibody. In early pre-dentin, collagen type I, laminin, tenascin and fibronectin were present. In late pre-dentin and dentin collagen type I was found in intertubular dentin and in the zone between enamel and dentin. The close relationship between collagen type I in dentin and fibronectin in immature enamel is interesting, as it may contribute to the stabilization of the amelodentinal interface. In dental pulp, collagen type IV and laminin were found in the endothelial basement membranes. Collagen type I and III, tenascin and fibronectin were localized to the mesenchymal intercellular matrix. The results of this study have supported the assumption that the lamina basalis ameloblastica is a basement membrane, and have lead to the suggestion that ameloblasts are producers of fibronectin or a fibronectin-like substance.

Ameloblasts↗

PubMed1994

Glycosylation pattern and enzyme activities in atrophic, angulated skeletal muscle fibres from ageing rats.

In this study enzyme activities and lectin binding patterns in skeletal muscle from very old rats were investigated in order to evaluate changes in enzyme activity or carbohydrate expression in senile muscle. Activities for adenosine triphosphatase (ATPase), succinic dehydrogenase, non-specific esterase and the binding pattern for 31 lectins were investigated in the soleus muscles from very old (36 months) and young (3 months) rats. In ageing muscles atrophic, angulated muscle fibres are frequent. In cryostat sections these fibres were mostly but not always type II defined by the myosin ATPase reaction; few showed a strong esterase activity. Some showed strong activity for succinic dehydrogenase while others were weakly reacting. A number of lectins strongly bound to the sarcoplasm in angulated fibres while the binding to normal fibres in both old and young rat muscle was much weaker or even absent. Preferential binding to the ageing, angulated fibres was seen with Aleuria aurentia, Galantus nivalis, Caragana abborecens, Triticum vulgaris, Maackia amurensis, Sambucus nigra, Phaseolus vulgaris erythroagglutinin, and Phaseolus coccineus. Samples of homogenized and centrifuged muscles were run by electrophoresis and the gels blotted to nitrocellulose paper. Subsequent lectin staining of the blots detected that two glycoproteins with molecular weights around 25,000 and 21,000 daltons were present in old muscle, but not in young. Aberrant or elevated expression of sarcoplasmic glycoconjugates is involved in ageing muscle atrophy.

Aging↗

PubMed1994

The protein effect on determination of DNA with Hoechst 33258.

The present study was designed to afford a critical review of the effect of proteins on the Hoechst 33258 method for determination of DNA of crude homogenates. A considerable effect of proteins on the fluorescence was observed when the concentration exceeded 100 micrograms BSA equivalent protein. Below that value, practically no effect of proteins was noted. We used proteinase K to remove the proteins, but dilution of homogenates could be used as well. Moreover, we found that the concentration of the fluorochrome should be between 1 microgram and 2 micrograms when microgram levels DNA are to be determined.

Animals↗