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PubMed1998

HPLC with fluorescence detection of urinary phenol, cresols and xylenols using 4-(4,5-diphenyl-1H-imidazol-2-yl)benzoyl chloride as a fluorescence labeling reagent.

A simple and sensitive HPLC method for the determination of phenolic compounds, i.e., phenol (Phe), cresols (Cres) and xylenols (Xyls), was developed. After a pre-column fluorescence derivatization of these compounds with 4-(4,5-diphenyl-1H-imidazol-2-yl)benzoyl chloride (DIB-Cl) at 60 degrees C for 30 min, 11 DIB derivatives were successfully separated within 50 min with an ODS column using CH3CN-H2O-CH3OH (25 + 22 + 53, v/v) as the eluent. The detection limits of DIB derivatives at a signal-to-noise ratio of 3 ranged from 0.15 to 1.09 microM (0.2-1.6 pmol per 20 microliters). The precision of the proposed method for both within- and between-day assays of free and total phenol related compounds was satisfactory (RSD < 9.5%). By the proposed method, Phe and p-Cre could be detected in normal urine samples, and the calculated concentrations of free Phe and p-Cre in unhydrolysed urine samples were 1.5 +/- 1.3 and 23.9 +/- 24.3 microM and those of total Phe and p-Cre in hydrolysed urine samples were 87.3 +/- 81.2 and 200.7 +/- 195.4 microM (n = 21), respectively.

Benzoates↗

PubMed2001

A case of adult T-cell leukaemia/lymphoma characterized by multiplex-fluorescence in situ hybridization, comparative genomic hybridization, fluorescence in situ hybridization and cytogenetics.

Adult T-cell leukaemia/lymphoma (ATLL) is a neoplasm of mature helper (CD4) T lymphocytes. Little is known, however, about the chromosome aberrations associated with the pathogenesis of this malignancy. Using molecular cytogenetic techniques we, therefore, investigated a 44-year-old man who had a 7-year history of ATLL with cutaneous involvement mimicking primary cutaneous T-cell lymphoma. Conventional cytogenetics revealed gross chromosomal changes with chromosome numbers ranging from 71 to 82. There were structural abnormalities of chromosomes 7 and 9, partial deletions of chromosomes 1, 3, 5 and 6, and loss of chromosomes 2, 4, 9, 11--14, 21 and 22. Multiplex-fluorescence in situ hybridization (M-FISH) identified two derivative chromosomes, der(6)t(6;7)(q16;q21) and der(7)t(6;7)(q16;q21)ins(6;12)(q2?;?), and a deletion of chromosome 1p. Conventional FISH confirmed the M-FISH findings. Comparative genomic hybridization of the blood revealed gains of DNA copy number at 1q12--25, 6p24--25, 9p23, 16p13--q13, 17q11--21, 19p13 and 20q13 and loss at 11p15 while lymph nodes showed gains at 3p22--24, 3q27--29, 7q36 and 15q26 and losses at 2p24--25, 2q37, 10p14--15, 11p15, 13q33--34 and 16p13.3. No DNA copy number changes were seen in a skin lesion. These results show the extent of genetic abnormalities within this malignancy.

Adult↗

PubMed2005

Delineation of a supernumerary marker chromosome utilizing a multimodal approach of G-banding, fluorescent in situ hybridization, confirmatory P1 artificial chromosome fluorescent in situ hybridization, and high-resolution comparative genomic hybridization.

We describe the structure of a supernumerary marker in a child who presented with a right atretic ear and multiple congenital anomalies. Using G-banding, fluorescent in situ hybridization (FISH), P1 artificial chromosome FISH and high-resolution comparative genomic hybridization (CGH), the marker was demonstrated to be a derivative chromosome resulting from malsegregation of a paternal 8;22 translocation: 47,XY, +der(22)t(8;22)(q24.1; q11.2). This case is noteworthy because the marker, while sharing similarities to der(22) in the Cat Eye syndrome (CES), also contains chromosome 8q material. This partial 8q trisomy confounds the diagnosis of CES associated with pure trisomy or pure tetrasomy 22q. The paternal translocation is noted with prolonged infertility and oligospermia, which again highlights the utility and necessity of chromosome analysis in this setting.

Abnormalities, Multiple↗

PubMed2002

Using fluorescence-activated cell sorting followed by fluorescence in situ hybridization to study lineage relationships: the 8;21 translocation is present in neutrophils but not monocytes in a patient with severe congenital neutropenia and a granulocyte colony-stimulating factor-responsive clonal abnormality.

UNLABELLED: Severe congenital neutropenia (Kostmann syndrome) is a disorder that presents in the neonatal period, but predisposes to leukemia later in life. This report describes a 4-y-old female with a history of severe congenital neutropenia, who developed a clonal abnormality associated with the translocation (7;21;8) (q32;q22;q22) (AML-1/ETO). She had circulating peripheral blasts and bone marrow blast counts as high as 64% when she received recombinant granulocyte colony-stimulating factor (rG-CSF). Her marrow blasts decreased to 4-20% when rG-CSF was discontinued. Fluorescence in situ hybridization analysis was performed on bone marrow cell populations sorted by flow cytometry to determine which cell populations had the AML-1/ETO translocation. The translocation was found in mature neutrophils and blasts, but not in monocytes, lymphocytes or stem cells. CONCLUSION: These findings suggest that the translocation occurred in a neutrophil progenitor, past the point in ontogeny where monocytes and neutrophils separate. The techniques described may be useful in understanding lineage relationships and leukemogenesis in other clonal abnormalities associated with myelodysplasia and leukemia.

Bone Marrow Transplantation↗

PubMedDate not supplied

Two-photon 3-D mapping of ex vivo human skin endogenous fluorescence species based on fluorescence emission spectra.

Spectral resolved tissue imaging has a broad range of biomedical applications such as the minimally invasive diagnosis of diseases and the study of wound healing and tissue engineering processes. Two-photon microscopy imaging of endogenous fluorescence has been shown to be a powerful method for the quantification of tissue structure and biochemistry. While two-photon excited autofluorescence is observed ubiquitously, the identities and distributions of endogenous fluorophores have not been completely characterized in most tissues. We develop an image-guided spectral analysis method to analyze the distribution of fluorophores in human skin from 3-D resolved two-photon images. We identify five factors that contribute to most of the luminescence signals from human skin. Luminescence species identified include tryptophan, NAD(P)H, melanin, and elastin, which are autofluorescent, and collagen that contributes to a second harmonic signal.

Equipment Design↗

PubMedDate not supplied

The role of the protein matrix in green fluorescent protein fluorescence.

In the ground state of the highly conjugated green fluorescent protein (GFP), the chromophore should be planar. However, numerous crystal structures of GFP and GFP-like proteins have been reported with slightly twisted chromophores. We have previously shown that the protein cavity surrounding the chromophore in wild-type GFP is not complementary with a planar chromophore. This study shows that the crystal structure of wild-type GFP is not an anomaly: most of the GFP and GFP-like proteins in the protein databank have a protein matrix that is not complementary with a planar chromophore. When the pi-conjugation across the ethylenic bridge of the chromophore is removed the protein matrix will significantly twist the freely rotating chromophore from the relatively planar structures found in the crystal structures. The possible consequences of this nonplanar deformation on the photophysics of GFP are discussed. A volume analysis of the cis-trans-isomerization of HBDI, a GFP chromophore model compound, reveals that its hula-twist motion is volume conserving. This means that, if the GFP chromophore or GFP chromophore model compounds undergo a cis-trans-isomerization in a volume-constricting medium, such as a protein matrix or viscous liquid, it will probably isomerize by means of a HT-type motion.

Biosensing Techniques↗

PubMed1979

Separation of mouse T cell subsets by a fluorescent activated cell sorter using fluorescence-labeled peanut agglutinin.

Nylon non-adherent spleen T cells obtained from concanavalin A-injected mice were labeled with fluorescein-labeled peanut agglutinin and separated into peanut agglutinin-positive (PNA+) cells and peanut agglutinin-negative (PNA-) cells by a fluorescent activated cell sorter. PNA+ cells were found to exert marked suppressive effect on primary anti-sheep red blood cells antibody response, but PNA- cells did not affect the antibody response. From these results and from the sugar-binding specificity of PNA, suppressor T cells are supposed to possess abundant galactosyl residues exposed on the cell surface, which are not masked by sialyl residues.

Animals↗

PubMed1999

[ANS fluorescence. II. Use of the method of fluorescent probe decay for studying the structural transformations in globular proteins].

Changes in ANS fluorescence decay parameters induced by the interaction of the probe with proteins have been investigated. The existence of at least two different modes of interactions between the ANS and protein was established. The interactions of the first type are connected with binding of an ANS molecule with the surface of a protein molecule. In this case ANS molecules are well acceptable for a solvent. The interactions of the second type are characteristic of the protein-embedded ANS molecules. The decay time values of the second type complexes change considerably (> 1.5-fold) during the protein molecule transformation into the molten globule-like conformation. The molecular model explaining such a behaviour is suggested.

Anilino Naphthalenesulfonates↗

PubMedDate not supplied

[Effect of treated with weak magnetic field aqueous salt solutions on the intrinsic fluorescence of bovine serum albumin. Isolation from solutions and partial characterization of the biologically active fluorescing fraction].

It was shown that water with additions of Ca2+, Na+, K+ and Cl- ions preliminarily treated with weak combined constant (42 microT) and low-frequency alternating (0.06 microT) magnetic fields affects the intrinsic fluorescence of bovine serum albumin, the magnitude of the effect being dependent on the frequency of the alternating field and ionic composition of the aqueous salt solution. A practically complete transfer of the effect through a small portion of the solution treated with magnetic fields was revealed. It was also found that after magnetic treatment, the solution contains a rather large (molecular mass 700-900 D) and stable molecular associate, which possesses, at least partially, the properties and characteristics inherent in the whole solution that were as acquired as a result of magnetic treatment.

Calcium Chloride↗

PubMedDate not supplied

[Fluorescent substrates of proteolytic enzymes with internal quenching of fluorescence].

The design and application of a successfully developed type of fluorogenic substrates for proteolytic enzymes are discussed. Proteolytic enzymes play a key role not only in the regulation of cellular protein turnover but also in the control of many other physiological functions. Substrates consisting of the appropriate peptides with chromophore or fluorophore at the C-termini are often employed to detect protease activity. Proteases exhibit the primary specificity to the particular amino acid residue and hydrolyze the following peptide bond. However, it is impossible to investigate the specific recognition of the other side of a scissible bond. Intramolecularly quenched fluorescent substrates are very useful instruments for research and medical purpose due to their high sensitivity and selectivity to proteases.

Amino Acid Sequence↗

PubMed2003