Trennung der intestinalen β-Galactosidasen beim lactose-toleranten Erwachsenen durch Ultrazentrifugation im Dichtegradienten

Zusammenfassung Die intestinalen-Galactosidasen von 4 lactose-toleranten, erwachsenen Mitteleuropäern wurden im Saugbiopsie-Gewebe nach Solubilisierung mit Triton X-100 in einem linearen Mannitol-Gradienten (5–20%) auf der Ultrazentrifuge bei 4°C und 44000 U/min getrennt. Bei 12stündiger Zentrifugation fanden sich 3 Fraktionen, von denen die beiden schneller sedimenticrenden Lactose spalten. Alle 3 Fraktionen hydrolysieren p-Nitrophenyl--Galactosid. Die 3 isolierten-Galactosidasen entsprechen wahrscheinlich der neutralen Bürstensaum-Lactase, der sauren lysosomalen Lactase und einer cytoplasmatischen Hetero--Galactosidase.     

Phenotypic and epistatic grouping of hypo- and hyper-rec mus mutants in Aspergillus

The mutants musK to musS of Aspergillus nidulans are sensitive to methyl-methanesulfonate (MMS) and several of them are meiotic-defective and alter mitotic recombination frequencies. All were found to be cross-sensitive to 4-nitro-quinoline-N-oxide (4-NQO) but unexpectedly none of them was hypersensitive to -rays and few to UV light. Double mus;uvs mutants were constructed to test for interactions with uvs mutations of the four epistatic groups of Aspergillus, UvsF, UvsC, UvsI, and UvsB. All meiotic-defective mus mutations caused some lethal interactions, usually with uvsF. None of them showed epistasis with UvsF or UvsB group mutants and one, musO, may represent a new group. Three mus mutations that affect recombination were assigned to the UvsC group, namely musN and K, and also musL which is recombination-defective and closely resembles uvsC. While uvsC mutants are mutators and lack UV-mutagenesis, most mus mutants had no effects on mutation. Only musR, which appeared epistatic with uvsI, showed reduced UV-reversion frequencies similar to uvsI. The recombination-proficient mus mutants appeared to be epistatic with more than one group, but in several cases sensitivities were slight and overlaps insufficient to obtain corroborating results with MMS and 4-NQO.     

Diameter and flow velocity changes of feline small pulmonary vessels in response to sympathetic nerve stimulation

Using an X-ray television system, we measured directly changes in the internal diameter (ID), flow velocity, and volume flow of the small pulmonary vessels (100–500 m ID) in response to electrical sympathetic nerve stimulation (SNS) in anaesthetized cats before and after adrenergic receptor blockade. Flow velocity was obtained by measuring the distance that the leading edge of the contrast medium moved per 0.1 s in the small arteries. Volume flow was obtained from the product of flow velocity and cross-sectional area calculated from the ID of the small arteries. SNS was accolmplished with 10- to 15-V square-wave pulses of 2-ms duration at 20–30 Hz for 20-s periods. In response to SNS, arterial ID decreased significantly by 8–13% in the 200- to 500-m vessels but not in the 100- to 200-m vessels. In the veins, on the other hand, there was no significant ID decrease in any of the 100- to 500-m vessels. After -receptor blockade (phentolamine, 2 mg/kg i.V.), there were significant ID increases (4–9%) in the 100- to 500-m arteries in response to SNS, the maximum increases being in the 100- to 200-m arteries. After -blockade (propranolol, 2 mg/kg i.V.), the ID decrease due to SNS in the 200- to 500-m arteries was enhanced (24–27%) and, in addition, the 100- to 200-m arteries exhibited a significant ID decrease (18%). Combined and -blockade completely abolished the ID decrease due to SNS. In the veins, on the other hand, no ID change occurred even after - or -blockade. The results indicate that SNS selectively constricts 200- to 500-m arteries. The data suggests that SNS has -mediated vasoconstrictor and -mediated vasodilator effects on the 100- to 500-m arteries and that the ID response pattern to SNS depends chiefly on the balance between -mediated vasoconstriction and -mediated vasodilation. Associated with the ID decrease due to SNS, flow velocity was increased by 21%. However, SNS did not affect volume flow, because the increase in velocity was compensated by the reduction in the cross-sectional area (due to the decreased ID).     

Application of the polymerase chain reaction to study the M protein(-like) gene family in beta-hemolytic streptococci

Evaluation of homologous regions of published M protein (emm) gene sequences from group A streptococci (GAS; Streptococcus pyogenes) was used to design three primer pairs for polymerase chain reaction (PCR) and three oligonucleotide probe sequences internal to the amplified products. One set of primers and corresponding probe should detect and lead to amplification of emm(-like) genes of virtually every type (all M), another (SOR^-M) should only amplify emm(-like) genes from GAS negative for serum opacity reaction (SOR) and the third (SOR⁺M) should expand only emm(-like) genes from SOR⁺ GAS. Using the all M primer pair for PCR on the genomic DNA from GAS of 29 different M types as well as from a group C and a group G streptococcal isolate, DNA fragments within the expected size range were amplified in every assay. All PCR products reacted with the all M probe. Related sequences were not detected in genomic DNA of an S. agalactiae and an Enterococcus faecalis isolate. Applying the SOR^-M and SOR⁺M primers to identical assays led to mutually exclusive amplification products. The SOR⁺M and SOR⁺M probes hybridized only to their corresponding products. Exceptions to this exclusivity were the SOR⁺ GAS of M types 3, 8, 27, 34, 42, 67, and 69, which consistently reacted only with the SOR⁺M primer/probe set. Analysis of sequence data from the amplified emm(-like) 2, 3, 18, and 19 genes revealed interesting specific features such as conserved gaps in the C-terminal sequence regions from SOR⁺ and the exceptional SOR^- GAS strains. These data indicate the existence of a subgroup of strains among SOR^- GAS and may advance our understanding of phylogenetic relationship between different serotypes of GAS.     

Down-regulation of tumor necrosis factor α activity by acute ethanol treatment in human peripheral blood monocytes

As the most commonly used drug that can modulate both metabolic and immune pathways, ethanol is evaluated in this report as a regulator of tumor necrosis factor (TNF) production in human peripheral blood monocytes (M) in combination with a variety of stimuli. While acute ethanol treatment did not induce TNF in M, it was a potent down-regulator of M TNF production whether induced by the combination of interferon- plus muramyl dipeptide (MDP) (P<0.001), lipopolysaccharide (LPS) alone (P<0.01), or interferon- plus LPS. Down-regulation of M TNF by ethanol was dose dependent and statistically significant in the biologically relevant, 25–150 mM, ethanol concentration range. We also demonstrate that these ethanol concentrations did not affect M viability. TNF down-regulation by ethanol was most effective when ethanol was administered 4 hr prior to MDP stimulation; however, it was also effective—though to a lesser extent—if it was added at the time of MDP stimulation. Furthermore, ethanol also down-regulated TNF production of thein vivo preactivated M of trauma patients, which produce hyperelevated levels of TNF. We have previously shown that the majority of posttrauma elevated M TNF is produced by the M subpopulation expressing high-affinity type I Fc receptors (FcRI). When the FcRI cross-linking-stimulated M subpopulation was treated with acute ethanol, TNF production was suppressed again both inin vivo preactivated M of trauma patients and in M of normal controls. In experiments utilizing cyclooxygenase inhibitor, we also demonstrate that ethanol has a direct, prostaglandin E₂-independent, effect on M TNF production. These results demonstrate that acute ethanol exposure has the potential to down-regulate M production of TNF significantly regardless of the TNF-inducing stimulus. Decreased capacity of M to produce TNF might, therefore, contribute to the immunological and metabolic abnormalities described after ethanol uptake.     

Factors influencing serum neopterin and β2-microglobulin levels in a healthy diverse population

Sera and questionnaire data from a population-based random sample of healthy adults was used to evaluate factors influencing neopterin and 2-microglobulin (2m) values. Both neopterin and 2m levels increased with age and were higher among white than blacks (mean values for whites and blacks: neopterin, 5.06 vs 4.49 nmol/L; 2m, 1.36 vs 1.28 mg/L). Gender differences were noted for 2m but not neopterin values (2m males vs females: 1.37 vs 1.29 mg/L). Neopterin values were lower among current smokers than among nonsmokers (4.32 vs 5.16 nmol/L) and were higher among users of antihistamines (5.46 among users vs 4.65 nmol/L among nonusers). Neopterin and 2m were correlated in this healthy adult population (adjustedr=0.53,P=0.001), yet no other interrelationships with numerous biologic markers except between 2m and serum-soluble interleukin-2 receptor levels (adjustedr=.41,P=0.05) were observed. These findings provide important baseline information to consider before planning or evaluating studies utilizing neopterin or 2m levels.     

Kinetic and pharmacological properties of high- and low-threshold calcium channels in primary cultures of rat hippocampal neurons

The kinetic, permeability and pharmacological properties of Ca currents were investigated in primary cultures of rat hippocampal neurons. The low-voltage-activated (LVA) Ca current turned on positive to –60mV and fully inactivated in a voltage-dependent way. This current was depressed by nickel (Ni, 40 M) and amiloride (500 M) and was insensitive to -conotoxin (-CgTx) (4 M) and to the Ca agonist Bay K 8644 (5 M). The high-voltage-activated (HVA) Ca current turned on positive to –40 mV and inactivated slowly and incompletely. This current was much less sensitive than the LVA current to Ni and amiloride but more sensitive to cadmium. CgTx blocked only partially this current (about 50%) in an irreversible way. Bay K 8644 had a clear agonistic action almost exclusively on the -CgTx-resistant HVA current component. The present results suggest that the HVA channels, quite homogeneous for their kinetic properties and sensitivity to holding potentials, can be pharmacologically separated in two classes: (i) -CgTx-sensitive and Bay-K-8644-insensitive (-S/BK-I) and (ii) -CgTx-insensitive and Bay-K-8644-sensitive (-I/BK-S), the latter displaying a stronger Cadependent inactivation.     

Characterization and localization of α-connectin (titin 1): An elastic protein isolated from rabbit skeletal muscle

Summary A simplified procedure to isolate-connectin (titin 1, TI), a gigantic elastic protein, from rabbit skeletal muscle is described. A rapid column chromatography step to concentrate-connectin is introduced. Separation of-connectin from-connectin is introduced. Separation of-connectin from-connectin (titin 2, TII) in the presence of 4 M urea at pH 7.0 did not cause any change in the secondary structure of-connectin as judged by circular dichroic spectra. Ultraviolet absorption spectra and the amino acid composition of-connectin (MW, approximately 3×10⁶) were similar to those of its proteolytic product,-connectin (MW, approximately 2×10⁶). Circular dichroic spectra suggested that both- and-connectin consist of 60%-sheet and 30%-turn. It thus appears that the whole elastic filament of connectin has a folded-strand structure. Proteolysis of-connectin by calpain resulted in formation of-connectin and smaller peptides. The-connectin interacted with both myosin and actin filaments similarly to-connectin. Polyclonal antibodies raised against 1200 kDa peptides obtained from aged rabbit skeletal myofibrils reacted with-connectin (titin 1, TI) but only weakly with-connectin (titin 2, TII) in rabbit skeletal muscle. Immunoelectron microscopy and indirect immunofluorescence microscopy revealed that the antibodies bound at the Z-line and at the epitope regions in the I-band near the binding site of a monoclonal antibody SMI whose position depends on sarcomere length. It thus appears that-connectin extends from the edge of M-line to the above epitope region in the I-band.     

Altered synthesis of interferon-γ and expression of interferon-γ receptor by peripheral blood mononuclear cells from patients with IgA nephropathy and non-IgA proliferative glomerulonephritis

Previously we reported disease-specific interaction between interferon- (IFN-) and interleukin-4 (IL-4) in patients with IgA nephropathy (IgAN), suggesting the existence of unusual T cell behavior in this disease. In the present study, we investigated characteristic synthesis of interferon- (IFN-) and expression of IFN- receptor (IFN-R) in the peripheral blood mononuclear cells (PBMC) from patients with IgAN and other chronic proliferative glomerulonephritis (PGN). Heparinized peripheral blood samples were obtained from 38 patients with chronic mesangial proliferative glomerulonephritis (CGN; including 24 with IgA nephropathy) and 20 healthy controls. PBMC were isolated by gradient centrifugation and fragments were cultured in Iscove's modified Dulbecco's medium (IMDM) supplemented with 10% fetal calf serum (FCS) for 72 hr. IFN- concentrations in supernatants were evaluated by the enzyme-linked immunosorbent assay (ELISA). Other parts of PBMC pellets were reacted with anti-human IFN-R monoclonal antibody and FITC-labeled anti-mouse second antibody for analysis of IFN-R expression on these cells by FACScan. The remaining PBMC were fractionated into CD4⁺ T cells, CD8⁺ T cells, B cells, NK, cells and macrophages using the MACS cell sorting system. The isolated cells were evaluated for IFN- or IFN-R mRNA expression by the semiquantitative RT-PCR method.In vitro IFN- synthesis was enhanced in patients with CGN, and NK cells were revealed to be responsible for such enhancement. On the other hand, the expression of IFN-R on macrophages was suppressed in CGN patients. These results suggest that impairment of regulation of the IFN- system might be involved in the development of CGN.     

Influence of adoptively transferred thioglycollate-elicited peritoneal macrophages on metastasis formation in mice with depressed or stimulated NK activity

The effect of thioglycollate-elicited macrophages (TG-M) on natural killer (NK)-cell activity and metastases formation in mice was investigated. Intravenously (i.v.) inoculated TG-M inhibited spleen NK activity of normal mice and abrogated polyinosinic: polycytidylic (poly IC) induced augmentation of NK cell function. TG-M also inhibited the clearance of i.v.-injected radiolabeled B16 melanoma cells from the lungs of normal or poly IC stimulated mice. Formation of experimental B16 melanoma metastases was dramatically increased in mice pretreated with TG-M. Administration of TG-M increased metatasis formation to a greater extent than anti-asialo GM₁ serum, while anti-asGM₁ serum was more efficient than TG-M in depressing spleen NK cell activity. When mice with low NK reactivity (beige mice or mice treated with anti-asialo GM₁ serum) were inoculated with TG-M, there was a substantial additive augmenting effect on metastasis formation in the lungs. Treatment with poly IC elevated NK-cell activity and had profound antimetastatic effects in normal but not in TG-M pretreated mice. The metastasis augmenting effect of TG-M was fully expressed in poly IC-treated mice as well as in athymic nude mice. Inoculation of proteose peptone-elicited macrophages (PM), unlike TG-M, did not depress NK activity or augment metastasis formation in normal or poly IC-treated mice. However, since the inhibition of NK activity in TG-M-treated mice was relatively weak, and a substantial additional increase in metastases was observed in NK-depressed mice after transfusion of TG-M, it seems unlikely that the TG-M-induced inhibition of NK reactivity is entirely responsible for the augmented formation of metastases. Further studies revealed that i.v. inoculation of TG-M, but not PM, induced intravascular inflammatory reactions, and damage to endothelial cells and basement membrane of the lung vasculature. These reactions may contribute to increased tumor cell extravasation and metastasis formation in mice pretreated with TG-M.     

Combination therapy of cyclosporine with steroid inhibits γ-interferon and interleukin-1 gene expression at the level of mRNA synthesisin vivo

The present study examined the effect of cyclosporine (CsA) administered with steroidin vivo on the capacity of peripheral blood mononuclear cells (PBMC) from kidney transplant recipients to generate cytokines and their gene expression at the level of messenger RNA (mRNA). PBMC from CsA-prednisolone (Pred)-treated recipients displayed 66.9% inhibition (54.3±12.4 IU/ml;N=42;P<0.01) of -interferon (-IFN) production compared with normal individuals (134.6±18.6 IU/ml;N=23). Azathioprine (Az)-Pred-treated recipients displayed significantly less inhibition of -IFN generation (96.0±16.1 IU/ml;N=22;P<0.05) than CsA-treated patients. Macrophages (m) from CsA-Pred-treated recipients displayed 60.0% inhibition (5.1±0.7 U/ml;N=20;P<0.01) of interleukin-1 (IL-1) production compared with normal individuals (13.0±2.9 U/ml;N=21). These results were confirmed by the experiments using cDNA probe for -IFN or IL-1 (, ). High levels of -IFN mRNA in phytohemagglutinin (PHA)-stimulated PBMC or IL-1() mRNA in lipopolysaccharide (LPS)-stimulated m were present in normal individuals but not in CsA-treated recipients as judged by hybridization to a cloned human -IFN or IL-1() cDNA probe. These studies demonstrated that combination therapy of CsA with steroid inhibits both -IFN and IL-1 gene expression at the level of mRNA at physiological concentrations.     

Serum interferon activity in inflammatory bowel disease

Serum interferon (IFN) of-class was studied in 64 consecutive patients, 26 with Crohn's disease, 38 with ulcerative colitis, and in 34 healthy volunteers. Detectable IFN- in 10 patients was associated with a moderate to severe activity of chronic inflammatory bowel disease (CIBD). However, 19 of 28 patients (68%) with activity in their disease did not have elevated IFN- levels. The three groups, ulcerative colitis, Crohn's disease, and healthy volunteers did not reveal any statistically significant difference in serum IFN-, as four of 34 healthy controls without intercurrent infections had elevated levels as well. Possible effects of, , and classes of IFN on endogenous arachidonic acid (AA) release and metabolism in human neutrophils was investigated in a substudy. IFN- caused a dose-dependent release of AA from phospholipids and metabolism of a modest fraction of leukotriene B₄ (LTB₄) and 5-hydroxyeicosatetraenoic acid (5-HETE) at doses reaching a maximum of 100 IU/ml. IFN of the and classes did not exert such effects. Addition of complement 5a to cells activated by IFN- caused induction of increased 5-li-poxygenase activity with unchanged release of AA. As only 16% of all CIBD patients had elevated IFN- levels as compared to 12% among the group of healthy volunteers, IFN- does not seem to be of importance for the perpetuation of the inflammatory reaction in ulcerative colitis or Crohn's disease, and other factors may therefore be responsible for activation of the inflammatory cells to production of LTB₄ and 5-HETE.     

Hepatozoon canis: Size measurement of the gametocyte using image analysis technology

This study presents definitive dimensions of the gametocyteHepatozoon canis using image analysis technology. The mean length and width of the gametocyte was found to be 11.42 ,m and 5.39 m, respectively. The mean area of the gametocyte was found to be 45.88 m². The average perimeter of the gametocyte was 28.92 m. The results compare well with some previous measurements using light-microscopic techniques.     

Über zwei neue anabol wirksame Steroide

Zusammenfassung 1-Methyl- ¹-androsten-17-ol-3-on-17-acetat ist unter zahlreichen von uns geprüften das zur Zeit am stärksten wirksame anabole Steroid mit der relativ geringsten androgenen Nebenwirkung. Die Beeinflussung des Cyclus bei der normalen Ratte ist gering. Eine Überlegenheit gegenüber Testosteron-17-propionat tritt am chronisch dihydrotachysterin-vergifteten Tier deutlich hervor, die sich neben der Erhaltung des Körpergewichts in der Verhütung unphysiologischer Ca-Ablagerungen manifestiert. Die anti-ulcerogene Wirkung des 1-Methyl- ¹-androsten-17-ol-3-on-17-acctat ist gleich der des Testosteron-17-propionat; hier liegt die Überlegenheit des 1-Methyl- ¹-androsten-17-ol-3-on-17-acetats in seiner schwächeren androgenen Nebenwirkung.Das 1-Methyl- ¹-androsten-17-ol-3-on-17-önanthat zeigt besonders starke und langdauernde anabole Wirkung bei sehr geringer und wesentlich kürzerer androgener Wirksamkeit.     

Direct observation of sexual competition inDrosophila melanogaster: The mutantWhite in competition with other genotypes

When two types ofDrosophila are in competition, the frequency dependence of mating success routinely is measured in our laboratory by direct observation of mating pairs in Elens-Wattiaux observation chambers. The present experiments concern white mutants (used here as a reference standard) in competition with three other genotypes: wild-type Canton S, giantgt w ^a , and giantgt ^13z /w ⁺ Y/C(1)Dx, y f. From the present observations, the frequency dependence of mating success seems a very common phenomenon: a rare-type sexual advantage exists for females and for males. Sexual activity of male and femalewhite mutants is significantly higher when food is present in the mating chamber. Males of the other strains also are more active in the presence of food. Homogamic matings are more frequent amonggt ^13z /w ⁺ Y/C(1)Dx, y f flies.     

Immunohistochemical studies of human FSH producing pituitary adenomas

Summary Ten FSH producing pituitary adenomas were studied immunohistochemically. 9 cases were in males, and 7 showed elevated serum FSH levels. Immunohistochemically, all cases showed the presence of -subunit and FSH- subunits in many tumour cells. These two subunits were frequently colocalized in the same cells. However, the expression of LH- subunit was extremely low (1 of 10 cases exhibiting occasional LH- positive tumour cells), although it has been reported that FSH- and LH- subunits are colocalized in the same cells of the normal adult pituitary gland. Immunoelectron microscopically, -subunits and FSH- were present in the secretory granules and suggested the co-release of subunits or secretion of combined form of FSH. In 7 cases, TSH- was positive, and in some cases, TSH- was colocalized in the same tumour cells which contained -subunit and FSH- subunit. A few cases also demonstrated immunoreactivity for PRL and ACTH. Our immunohistochemical studies suggest that FSH adenomas are multihormonal and that there is abnormal gene expression in FSH cells with loss of LH- appearance and co-expression of TSH-.     

Quantitative Immunglobulinbestimmungen im Seminalplasma

Zusammenfassung Es wird über quantitative Immunglobulinbestimmungen im Seminalplasma berichtet.G,A undM sind in unterschiedlicher Menge nachweisbar, wobei jedoch keine Abhängigkeit von den verschiedenen Spermaqualitäten besteht.

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Summary Quantitative estimations of immunoglobulins in seminal plasma were performed.G,A andM are present in various amounts, but there is no dependance in regard to the quality of semen.

     

ω-Grammotoxin blocks action-potential-induced Ca2+ influx and whole-cell Ca2+ current in rat dorsal-root ganglion neurons

Field-potential stimulation of rat dorsal-root ganglion (DRG) neurons evoked action-potential-mediated transient increases in intracellular free calcium concentration ([Ca²⁺]_i) as measured by indo-1-based microfluorimetry. Field-potential-evoked [Ca²⁺]_i transients were abolished by tetrodotoxin, and their dependence on stimulus intensity exhibited an abrupt threshold. -Conotoxin GVIA (-CgTx, 100 nM) inhibited action-potential-mediated Ca²⁺ influx by 79%, while nitrendipine (1 M) had little effect. -Grammotoxin SIA (-GsTx, 267 nM), a peptide toxin purified from the venom of the tarantula spider, Grammostola spatulata, blocked action-potential-mediated Ca²⁺ influx as effectively as did -CgTx, suggesting that -GsTx blocks N-type Ca²⁺ channels. In contrast to block by -CgTx, the block produced by -GsTx reversed upon washout of the peptide. -GsTx (270 nM) blocked 80%, and -CgTx (1 M) blocked 64%, of whole-cell Ca²⁺ current (I _Ca) elicited by step depolarization to 0 mV from a holding potential of –80 mV. -GsTx completely occluded inhibition of I _Ca by -CgTx. However, when applied after -CgTx, -GsTx produced an additional inhibition of 27%, indicating that -GsTx also blocked a non-N-type Ca²⁺ channel. BayK8644 (1 M) elicited an increase in I _Ca in the presence of maximally effective concentrations of -GsTx, suggesting that -GsTx does not block L-type channels. Thus, -GsTx displays a selectivity for Ca²⁺ channel subtypes which should prove useful for studying Ca²⁺ channels and Ca²⁺-channel-mediated processes.     

Phenotypic differences between induced and spontaneous 2μ-plasmid-free segregants of Saccharomyces cerevisiae

Summary The maximum specific growth rates (_max) of 2 -plasmid-free ([cir°]) segregants of three haploid and one diploid strain of Saccharomyces cerevisiae have been determined and compared with the _max of their 2 -plasmid-containing ([cir ⁺]) progenitors. Two classes of [cir°] strains have been examined: those induced by transformation with a 2 -based recombinant plasmid according to the method of Dobson et al. (1980) and those isolated as spontaneous [cir°] segregants from glucose-limited continuous cultures. The _max of the spontaneous [cir°] segregants was not found to differ significantly from that of their [cir ⁺] parents. In all cases, however, the induced [cir°] strains had a _max which was significantly less than that of their [cir ⁺] counterparts. This effect was particularly marked in the case of the diploid strain where a 34% reduction in _max was observed. The implications of these results are discussed in terms of the effect of the transformation process on host yeast cells.     

A comparative immunohistological study of cerebellar enolases

Summary A comparative immunohistological study of the neurone-specific enolase and enolase, demonstrates the exclusive neuronal localization of enolase and its absence from glial cells. In contrast, enolase is located in astroglial cells. The validity of enolase as a neuronal marker and enolase as an astrocytic marker, is confirmed both by a double labelling technique, using antibodies to and to revealed with fluorescence or peroxidase in the same tissue sections, and by immunoelectronmicroscopy.