Abstract: The invention relates to the whole-body external optical imaging of gene expression. Specifically, methods for whole-body external optical imaging of gene expression and methods for evaluating a candidate protocol or drug for treating diseases or disorders using a fluorophore operatively linked to the promoter of a gene and external optical imaging are provided herein. Methods to screen for substances or genes that regulate target promoters are also provided.

Abstract: Compositions and methods which ameliorate the virulence of bacterial infection are described wherein the active ingredient modulates transmethylation reactions in bacterial cells. Particularly useful compounds are inhibitors of S-adenosyl methionine synthetase (SAMS), of S-adenosyl homocysteine hydrolase (SAHH) and of transmethylases.

Abstract: An in vitro model system for viral infection is comprised of a tissue block from adult tonsil or lymph node supported on a matrix which is flexible and porous and wherin the supported tissue block is cultured in a medium whose surface is congruent with the tissue block/matrix interface. The histoculture system can be used to screen for antiviral drugs and to monitor the course of viral diseases.

Abstract: An in vitro model system for viral infection and for immune response is comprised of a tissue block from adult tonsil or lymph node supported on a matrix which is flexible and porous and wherein the supported tissue block is cultured in a medium whose surface is congruent with the tissue block/matrix interface. The histoculture system can be used to screen for antiviral drugs, to monitor the course of viral diseases, and to monitor an immune response to antigen stimulation.

Abstract: A depletion method to inhibit tumor growth includes introducing a viral expression system capable of expressing methioninase or a fusion protein containing methioninase into a tumor contained in a vertebrate subject or cells thereof. The fusion protein may contain a fluorescent protein to permit monitoring of the completeness of the depletion method. The fusion protein can be used in vivo as well as in vitro screening protocols which employ the viral expression system. The expression system includes control sequences and means to integrate the nucleotide sequence into the genome of a host cell for expression. The method may also include treating the cells with isolated methioninase and/or with a therapeutic cell. The depletion method can be with other known therapies, such as maintaining the animal having the tumor on a methionine depleted diet.

Abstract: Homocysteinase enzymes are disclosed which have sufficient specificity for homocysteine, as compared to cysteine that hydrogen sulfide can be used as a measure of homocysteine in a biological fluid even in the presence of substantial amounts of cysteine, exceeding the level of homocysteine. The enzyme of desired specificity can be readily prepared by mutation and screening of naturally occurring homocysteinases or by constructing chimeric forms.

Abstract: The construction of multicopy expression systems for methioninase are disclosed. The higher expression systems employ a promoter operably linked to two or more copies of a tandemly repeated methioninase encoding sequence. Such multicopy expression systems were found to produce unexpectedly high levels of methioninase when expressed in an appropriate host.

Abstract: An improved synthesis of N,N,-disubstituted-p-phenylene-diamine (DSPDA) preferably di-n-butyl-p-phenylenediamine is disclosed. The improvement rests on the discovery that DSPDA and its salts are extremely sensitive to oxygen so that once the product DSPDA or its salt is formed, oxygen must be excluded from all manipulations.

Abstract: Enzymatic methods to determine the concentration of pyridoxal 5′-phosphate (PLP) in biological fluids are described. The methods of the invention are useful to assess risk for cardiovascular disease. The assay can be a homogeneous assay using the ability of PLP to function as a co-enzyme for homocysteinase and related enzymes and measuring the products of the reaction preferably spectrophotometrically. The invention also includes improvements in sensitivity of assays for measuring hydrogen sulfide production by measuring fluorescence as opposed to absorbance of the oxidized product of H2S with N,N-dialkyl p-phenylene diamine.

Abstract: A composition and method for treating disorders related to tyrosinase gene expression and melanin biosynthesis is disclosed. The composition comprises a tyrosinase encoding nucleotide sequence and an ORF-438 encoding nucleotide sequence derived from Streptomyces, adapted for expression in mammalian cells. Also disclosed is a model system for evaluating agents that affect pigmentation.

Abstract: A method to assess the level of folate in a biological sample comprises: providing said sample with glycine N-methyltransferase (GMT) and with an excess of S-adenosyl methionine (SAM) and of glycine; providing a control which contains no folate with said GMT and excess SAM and glycine in comparable amounts to those provided to the sample; and comparing the concentration of at least one product formed in the sample with the concentrations of said product formed in the control, whereby the difference in levels of said product in the sample as compared to the control is directly proportional to the level of folate in the sample.

Abstract: The present invention describes a method for targeted and specific delivery of beneficial compounds, including hair dyes, melanin, proteins, and nucleic acids for gene therapy, to hair follicle cells using liposomes encapsulating the beneficial compound. Particularly preferred methods describe delivery of hair dyes, melanin or tyrosinase to the hair follicle for the purpose of improving hair color or condition, the delivery of compounds which prevent alopecia or stimulate hair growth, either by encapsulating a compound in liposomes, or by encapsulating a nucleic acid capable of expressing a protein in liposomes. Also described are liposome compositions for practicing the methods.

Abstract: A method to follow the progression of metastasis of a primary tumor, which method comprises removing fresh organ tissues from a vertebrate subject which has been modified to contain tumor cells that express GFP and observing the excised tissues for the presence of fluorescence is disclosed. The fluorescence can also be monitored by observing the tissues in situ. Vertebrate subjects which contain GFP producing tumors are useful models to study the mechanism of metastasis, as well as to evaluate candidate protocols and drugs. In addition, subjects already harboring tumors can be treated so as to modify the endogenous tumors to contain GFP. This permits clinical applications. Finally, by injecting a contrast dye into a subject harboring a GFP-labeled tumor, angiogenesis in the tumor can be observed directly.

Abstract: A method to follow the progression of metastasis of a primary tumor, which method comprises optionally removing fresh organ tissues from a vertebrate subject which has been modified to contain tumor cells that express GFP and observing the tissues in situ or observing the excised tissues for the presence of fluorescence is disclosed. Vertebrate subjects which contain GFP producing tumors are useful models to study the mechanism of metastasis. In addition, subjects already harboring tumors can be treated so as to modify the endogenous tumors to contain GFP. This permits clinical applications. Finally, by injecting a contrast dye into a subject harboring a GFP-labeled tumor, angiogenesis in the tumor can be observed directly.

Abstract: A method to follow the progression of metastasis of a primary tumor, which method comprises removing fresh organ tissues from a vertebrate subject which has been modified to contain tumor cells that express GFP and observing the excised tissues for the presence of fluorescence is disclosed. The fluorescence can also be monitored by observing the tissues in situ. Vertebrate subjects which contain GFP producing tumors are useful models to study the mechanism of metastasis. In addition, subjects already harboring tumors can be treated so as to modify the endogenous tumors to contain GFP. This permits clinical applications. Finally, by injecting a contrast dye into a subject harboring a GFP-labeled tumor, angiogenesis in the tumor can be observed directly.

Abstract: The present invention discloses the cloning of a methioninase-encoding nucleic acid molecule from Pseudomonas putida and the construction of high-level expression modules containing the methioninase-encoding nucleic acid molecule. The invention further provides expression modules that use the T7 RNA polymerase promoter to express the isolated methioninase-encoding nucleic acid molecules. Expression modules employing the T7 promoter were found to produce unexpectedly high levels of methioninase. The present invention further provides purification methods to obtain highly pure, endotoxin free methioninase, chemically modified forms of methioninase, crystallized methioninase and lyophilized methioninase preparations. The present invention further provides therapeutical methods using the disclosed recombinant methioninase preparations.

Abstract: A method to follow the progression of metastasis of a primary tumor, which method comprises removing fresh organ tissues from a vertebrate subject which has been modified to contain tumor cells that express GFP and observing the excised tissues for the presence of fluorescence is disclosed. Vertebrate subjects which contain GFP producing tumors are useful models to study the mechanism of metastasis.

Abstract: The present invention is directed to introduce a replacement pigment into the hair shaft through the hair follicle using a formulation of a replacement pigment in a liposomal composition.

Abstract: Novel enzymatic methods to determine the concentration of homocysteine in biological fluids are described. In a typical embodiment of the invention, the biological fluid sample is from a patient, and the methods of the invention are useful to assess risk for cardiovascular disease. The novel methods of the invention involve use of particular homocysteinase enzymes that permit the determination of homocysteine concentrations in biological samples without interference from the concentrations of cysteine and/or of methionine that are routinely present in such samples. There is also provided a diagnostic kit for use in determining the amount of homocysteine in a biological sample comprising (a) a homocysteinase having the aformentioned characteristics, and (b) at least one reagent capable of being used to determine the amount of product formed in the homocysteinase reaction.

Abstract: The present invention provides a methods and apparatuses for use in providing rapid, selective and effective targeted delivery of beneficial substances into hair follicles. The invention relies on the use of vibrational method of delivery of substances of different size from small molecules up to particles compatible with the diameter of hair strand into the hair bulb. The skin is subjected to the vibration of about 1 Hz to about 100 Hz frequency and 0.1 mm to about 10 mm amplitude, an amount which is sufficient to cause mechanical shifts in different directions of skin structure without damage. This results in follicle effective uptake of the tested substances such as carbon particles of different size as well as melanin. The methods and apparatuses of the present invention are particularly useful in conjunction with photosensitive agents and electromagnetic irradiation. Such a combination is used to target the delivery of the agent to within hair follicles and then subsequently activate the agent.