Ein chromatographisches Verfahren zur Bestimmung typenspezifischer Poliovirus-Antikörper mit 32P-markiertem Polioviru

1963 ◽  
Vol 18 (10) ◽  
pp. 798-808 ◽  
Author(s):  
Reiner Thomssen
Keyword(s):  
Aluminium Hydroxide ◽  
Virus Antibody ◽  
Poliovirus Type ◽  
Specific Antibodies ◽  
In Vitro Technique ◽  
Step Procedure ◽  
Highly Sensitive

A highly sensitive in-vitro technique for determination of homologous type-specific antibodies against polioviruses is described. 32P-labelled poliovirus type 1. strain Mahoney, purified by a three-step-procedure (15 000 rpm, aluminiumhydroxide. Ecteolacellulose) is mixed with homologous type-specific antiserum. After incubation the mixtures are adsorbed to aluminium hydroxide. Virus-antibody-com-plexes are more firmly bound to this adsorbent than virus without antibody: the distinguishing parameter is the concentration of divalent phosphate in the adsorption or the elution fluid. The firmness of binding of virus-antibody-complexes to aluminium hydroxide is further a function of the dilution of antiserum. This leads to a suitable end-point for reading antiserum titers. There is a good correlation between titers determined by this method and titers, determined by common neutralization tests. The method may be used to judge the virus-specifity of radioactivity after purification of 32P-labelled poliovirus.

DETERMINATION OF BLOOD CORTICOIDS USING THE IN VITRO RESIN UPTAKE OF 3H-PREDNISOLONE

1968 ◽  
Vol 57 (1) ◽  
pp. 23-32 ◽  
Author(s):  
Hironori Nakajima ◽  
Mitsunori Murala ◽  
Masumitsu Nakata ◽  
Takeshi Naruse ◽  
Seiji Kubo
Keyword(s):  
Thyroid Function Test ◽  
Normal Subjects ◽  
Adrenal Function ◽  
Function Test ◽  
Before And After ◽  
Adrenal Response ◽  
Suppression Test

ABSTRACT The in vitro resin uptake of 3H-prednisolone was used for the determination of blood cortisol after addition of radioactive prednisolone followed by Amberlite CG 400 Type 1 to the test serum, and incubation of the mixture. The radioactivity of the supernatant was compared before and after the addition of the resin. The principle of this method is similar to that of the 131I-triiodothyronine resin uptake for the thyroid function test. The tests for the specificity, reproducibility and sensitivity gave satisfactory results. The mean basal value ± SD of the 3H-prednisolone resin uptake was 35.3 ± 9.2% in normal subjects, and 27.1 ± 4.8% in pregnant women. This method was valid in various adrenal function tests, i. e. the adrenal circadian rhythm, corticotrophin (ACTH) test, dexamethasone suppression test and the adrenal response to lysine-8-vasopressin. It proved to be a sensitive indicator of the adrenal function. These results suggest that this method should be useful for a routine adrenal function test.

The glycoproteins C and G are equivalent target antigens for the determination of herpes simplex virus type 1-specific antibodies

2010 ◽  
Vol 166 (1-2) ◽  
pp. 42-47 ◽  
Author(s):  
Thomas Scheper ◽  
Sandra Saschenbrecker ◽  
Katja Steinhagen ◽  
Andreas Sauerbrei ◽  
Waltraud Suer ◽  
...  
Keyword(s):  
Herpes Simplex Virus ◽  
Herpes Simplex ◽  
Virus Type ◽  
Herpes Simplex Virus Type ◽  
Specific Antibodies ◽  
Simplex Virus

scholarly journals Biochemical and Genetic Studies of the Initiation of Human Rhinovirus 2 RNA Replication: Identification of a cis-Replicating Element in the Coding Sequence of 2Apro

2001 ◽  
Vol 75 (22) ◽  
pp. 10979-10990 ◽  
Author(s):  
Kinga Gerber ◽  
Eckard Wimmer ◽  
Aniko V. Paul
Keyword(s):  
Rna Replication ◽  
Human Rhinovirus ◽  
Viral Sequence ◽  
Coding Region ◽  
Terminal Protein ◽  
Genetic Studies ◽  
Poliovirus Type ◽  
Coding Sequence

ABSTRACT We have previously shown that the RNA polymerase 3Dpolof human rhinovirus 2 (HRV2) catalyzes the covalent linkage of UMP to the terminal protein (VPg) using poly(A) as a template (K. Gerber, E. Wimmer, and A. V. Paul, J. Virol. 75:10969–10978, 2001). The products of this in vitro reaction are VPgpU, VPgpUpU, and VPg-poly(U), the 5′ end of minus-strand RNA. In the present study we used an assay system developed for poliovirus 3Dpol (A. V. Paul, E. Rieder, D. W. Kim, J. H. van Boom, and E. Wimmer, J. Virol. 74: 10359–10370, 2000) to search for a viral sequence or structure in HRV2 RNA that would provide specificity to this reaction. We now show that a small hairpin in HRV2 RNA [cre(2A)], located in the coding sequence of 2Apro, serves as the primary template for HRV2 3Dpol in the uridylylation of HRV2 VPg, yielding VPgpU and VPgpUpU. The in vitro reaction is strongly stimulated by the addition of purified HRV2 3CDpro. Our analyses suggest that HRV2 3Dpol uses a “slide-back” mechanism during synthesis of the VPg-linked precursors. The corresponding cis- replicating RNA elements in the 2CATPase coding region of poliovirus type 1 Mahoney (I. Goodfellow, Y. Chaudhry, A. Richardson, J. Meredith, J. W. Almond, W. Barclay, and D. J. Evans, J. Virol. 74:4590–4600, 2000) and VP1 of HRV14 (K. L. McKnight and S. M. Lemon, RNA 4:1569–1584, 1998) can be functionally exchanged in the assay with cre(2A) of HRV2. Mutations of either the first or the second A in the conserved A1A2A3CA sequence in the loop of HRV2 cre(2A) abolished both viral growth and the RNA's ability to serve as a template in the in vitro VPg uridylylation reaction.

scholarly journals Utilization of a new gold/Schiff-base iron(iii) complex composite as a highly sensitive voltammetric sensor for determination of epinephrine in the presence of ascorbic acid

RSC Advances ◽  
2016 ◽  
Vol 6 (104) ◽  
pp. 101888-101899 ◽  
Author(s):  
Adam Gorczyński ◽  
Maciej Kubicki ◽  
Klaudia Szymkowiak ◽  
Teresa Łuczak ◽  
Violetta Patroniak
Keyword(s):  
Ascorbic Acid ◽  
Schiff Base ◽  
Schiff Base Complex ◽  
Voltammetric Sensor ◽  
Base Complex ◽  
Highly Sensitive

A new voltammetric sensor based on an iron(iii) Schiff-base complex/Au composite is synthesized and applied for the in vitro detection of epinephrine.

Inhibitory effects of 3-imino-5-phenyl-3H-1,2-dithiole on poliovirus type 1 replication in vitro

1990 ◽  
Vol 14 (4-5) ◽  
pp. 267-277 ◽  
Author(s):  
A. Garozzo ◽  
M.R. Pinizzotto ◽  
P.M. Furneri ◽  
D. Baratta ◽  
F. Guerrera ◽  
...  
Keyword(s):  
Inhibitory Effects ◽  
Poliovirus Type

scholarly journals Highly Sensitive Micellar Enhanced Spectrofluorimetric Method for Determination of Mirtazapine in Tablets and Human Urine: Application to In Vitro Drug Release and Content Uniformity Test

2016 ◽  
Vol 2016 ◽  
pp. 1-8 ◽  
Author(s):  
Hany W. Darwish ◽  
Ahmed H. Bakheit ◽  
Raed M. Alharbi
Keyword(s):  
Drug Release ◽  
Human Urine ◽  
Dosage Form ◽  
Content Uniformity ◽  
In Vitro Drug Release ◽  
Spectrofluorimetric Method ◽  
Highly Sensitive ◽  
Spiked Human Urine

A highly sensitive and simple micelle enhanced spectrofluorimetric method was developed for assaying mirtazapine (MRZ) in REMERON® tablets and spiked human urine directly without the need of derivatizing agent. The basis of the current procedure is the examination of the relative fluorescence intensity (RFI) of MRZ in sodium lauryl sulphate (SLS) micellar medium. The RFI of MRZ in water was enhanced markedly on addition of SLS. The RFI was measured at 403 nm after excitation at 320 nm. The fluorescence-concentration relationship was linear over the range 1–500 ng/mL, with lower detection limit of 0.399 ng/mL. The proposed method was successfully applied to the determination of MRZ in dosage form and spiked human urine. Recovery percentages of MRZ utilizing the current method were99.05±1.83,98.37±1.96, and100.41±2.61% for pure powder, pharmaceutical dosage form, and spiked human urine, respectively. The application of the proposed method was extended to test content uniformity and the in vitro drug release of REMERON tablets, according to USP guidelines.

scholarly journals Genetic and Biochemical Studies of Polioviruscis-Acting Replication Element cre in Relation to VPg Uridylylation

2000 ◽  
Vol 74 (22) ◽  
pp. 10371-10380 ◽  
Author(s):  
Elizabeth Rieder ◽  
Aniko V. Paul ◽  
Dong Wook Kim ◽  
Jacques H. van Boom ◽  
Eckard Wimmer
Keyword(s):  
Genome Replication ◽  
Coding Region ◽  
Stem Loop ◽  
Genetic Changes ◽  
Poliovirus Type ◽  
Type 3 ◽  
Type Sequence

ABSTRACT In addition to highly conserved stem-loop structures located in the 5′- and 3′-nontranslated regions, genome replication of picornaviruses requires cis-acting RNA elements located in the coding region (termed cre) (K. L. McKnight and S. M. Lemon, J. Virol. 70:1941–1952, 1996; P. E. Lobert, N. Escriou, J. Ruelle, and T. Michiels, Proc. Natl. Acad. Sci. USA 96:11560–11565, 1999; I. Goodfellow, Y. Chaudhry, A. Richardson, J. Meredith, J. W. Almond, W. Barclay, and D. J. Evans, J. Virol. 74:4590–4600, 2000). cre elements appear to be essential for minus-strand RNA synthesis by an as-yet-unknown mechanism. We have discovered that the cre element of poliovirus (mapping to the 2C coding region of poliovirus type 1; nucleotides 4444 to 4505 in 2C), which is homologous to thecre element of poliovirus type 3, is preferentially used as a template for the in vitro uridylylation of VPg catalyzed by 3Dpol in a reaction that is greatly stimulated by 3CDpro (A. V. Paul, E. Rieder, D. W. Kim, J. H. van Boom, and E. Wimmer, J. Virol. 74:10359–10370, 2000). Here we report a direct correlation between mutations that eliminate, or severely reduce, the in vitro VPg-uridylylation reaction and produce replication phenotypes in vivo. None of the genetic changes significantly influenced translation or polyprotein processing. A substitution mapping to the first A (A4472C) of a conservedAAACA sequence in the loop of PV-cre(2C) eliminated the ability of the cre RNA to serve as template for VPg uridylylation and abolished RNA infectivity. Mutagenesis of the second A (A4473C; AAACA) severely reduced the yield of VPgpUpU and RNA infectivity was restored only after reversion to the wild-type sequence. The effect of substitution of the third A (A4474G; AAACA) was less severe but reduced both VPg uridylylation and virus yield. Disruption of base pairing within the upper stem region of PV-cre(2C) also affected uridylylation of VPg. Virus derived from transcripts containing mutations in the stem was either viable or quasi-infectious.

scholarly journals Lymphocyte-based Determination of Susceptibility to Malignant Hyperthermia: A Pilot Study in Swine

2010 ◽  
Vol 113 (4) ◽  
pp. 917-924 ◽  
Author(s):  
Saiid Bina ◽  
John Capacchione ◽  
Sheila Muldoon ◽  
Munkhuu Bayarsaikhan ◽  
Rolf Bunger
Keyword(s):  
Malignant Hyperthermia ◽  
High Performance ◽  
Human Lymphocytes ◽  
Malignant Hyperthermia Susceptibility ◽  
Swine Model ◽  
Arterial Blood ◽  
Stimulation Of

Background Malignant hyperthermia susceptibility (MHS) is diagnosed by an invasive in vitro caffeine-halothane contracture test (CHCT) carried out on biopsied skeletal muscle tissue. We are presenting a novel blood test approach for malignant hyperthermia testing in a swine model. Our main aim was to determine whether adenosine production from lymphocytes after 4-chloro-m-cresol (4CmC) stimulation distinguishes homozygous swine carrying the Arg615Cys mutation in the ryanodine receptor type 1 (RyR1) gene (MHS swine) from normal swine. Methods Lymphocytes were isolated from arterial blood (40 ml) obtained from MHS (n = 7) and normal (n = 7) swine. Cells were suspended in Hank's balanced salt solution and treated with 4CmC (0-10 mm) at 37°C in the presence of adenosine deaminase inhibitor. After termination and purification of samples, aliquots (50 μl) were assayed for adenosine content using high performance liquid chromatography. Results Baseline adenosine levels before stimulating lymphocytes with 4CmC were 0.025 ± 0.004 and 0.041 ± 0.006 μm (mean ± SEM) in lymphocytes from normal and MHS swine, respectively (P = 0.125). Maximum responses were achieved at 1 mm 4CmC for both cell-line groups. Adenosine levels after stimulation with 4CmC (1 mm) were 0.185 ± 0.009 and 0.397 ± 0.038 μm in lymphocytes from normal and MHS swine, respectively (P = 0.0035). There was no overlap between adenosine levels in stimulated lymphocytes from MHS and normal swine. Conclusion 4CmC stimulation of porcine lymphocytes induces increased adenosine formation in MHS cells relative to those from normal swine; evaluation of adenosine formation in response to RyR1 agonists in human lymphocytes is needed.

scholarly journals Fluorescence-based determination of enzyme activity of recombinant CYP51b1 (sterol 14α-demethylase) with coumarin derivatives

2010 ◽  
Vol 56 (1) ◽  
pp. 132-137 ◽  
Author(s):  
N.A. Petushkova ◽  
A.V. Lisitsa ◽  
V.F. Pozdnev ◽  
I.I. Karuzina
Keyword(s):  
Enzyme Activity ◽  
Acetic Acid ◽  
Electron Donor ◽  
Model System ◽  
Coumarin Derivatives ◽  
Current Investigation ◽  
Soluble Model ◽  
Highly Sensitive

The current investigation was undertaken with the aim to carry out an in vitro evaluation of the ability of coumarin derivatives as probe substrates to predict the activity of CYP51b1. The results obtained indicate that 7-aminocoumarin-4-acetic acid (ACAC) can be used to determine the recombinant CYP51b1 activity. Determination of CYP51b1 activity with ACAC is based on the direct registration of fluorescence increasing at 30°C. It was found also that BMR in a simple soluble model system can be used as an electron donor for CYP51B1. Fluorescence-based assay is highly sensitive and can be used for the screening of sterol 14alpha-demethylase inhibitors.

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