bluuebunny/crossref_metadata_2025_split huggingface dataset:
Novel use of the BD FACS™ SPA to automate custom monoclonal antibody panel preparations for immunophenotyping
Authors: Kelliher Abigail S. , Parent David W. , Anderson David C. , Dorn Michelle E. , Hahn Jessica L. , Eapen Sara , Preffer Frederic I. 
Published: May 2005

AbstractBackgroundThe effective and accurate diagnosis of hematologic malignancies relies on flow cytometric immunophenotyping. Selected combinations of monoclonal antibodies (mAbs) arranged in multicolor panels allow for the accurate definition of normal and abnormal hematologic cell populations. The most time‐consuming and crucial step in the staining process involves dispensing combinations of multiple mAbs into their appropriate staining tubes. This step is prone to error, requires concentration and accuracy, and is dependent on technologist experience.The Becton Dickinson BioScience (BD) FACS™ Sample Prep Assistant (SPA) is touted as a breakthrough in automated in vitro diagnostic sample preparation. The SPA is designed to automate BD MultiTEST™ and BD TriTest lyse/no‐wash assays. However, because most cases in our laboratory require tedious application of unique four‐color mAb cocktails for leukemia and lymphoma testing, we wondered whether the SPA would be helpful in accurately dispensing these mixtures.MethodsThe mAb panels were prepared by the SPA in two separate timed runs and on separate days. Eleven specimens (nine from patients and two from normal volunteers) were split and stained with four‐color cocktails created by the SPA or manually. The percentage of positive (%P) cells and mean fluorescent intensity for each mAb pair were determined. These values were plotted against each other and correlation values were calculated. To quantitate timesaving in the laboratory, two technologists prepared individually the same mAb panels and were timed.ResultsThe correlation between the two methods was high; r2 was 0.988 for 158 %P antigen pairs; no bias between the manual and robotic methods was detected with the Wilcoxon rank test. Bland‐Altman analysis indicated no obvious relation between the difference and the mean of %P cells, suggesting that the SPA successfully dispensed antibodies for leukemia/lymphoma panels. The two methods may be interchangeable, although the limited sample size prohibits this conclusion from Bland‐Altman statistics alone. In addition, one possible error was detected in the SPA‐prepared panels. The SPA averaged 65 min/run, the experienced technologist 12.95 min/run, and the inexperienced technologist 54.9 min/run.ConclusionsSPA dispensing time was twice the average manual dispensing time; however, SPA use was completely automated and freed the technologist to perform other tasks. SPA use permitted preemptive preparation of mAb panels and thus streamlined processing; however, the cost of the assay and the amount of reagent waste increased. It is certain that software modifications by BD could decrease the SPA reagent dispense time and decrease the cost associated with reagent waste when the SPA is used in this novel fashion. © 2005 Wiley‐Liss, Inc.
Ultrastructural analysis of the smooth‐to‐striated transition zone in the developing mouse esophagus: Emphasis on apoptosis of smooth and origin and differentiation of striated muscle cells
Authors: Wörl Jürgen , Neuhuber Winfried L. 
Published: May 2005

AbstractThe exact mechanism of smooth‐to‐striated muscle conversion in the mouse esophagus is controversial. Smooth‐to‐striated muscle cell transdifferentiation vs. distinct differentiation pathways for both muscle types were proposed. Main arguments for transdifferentiation were the failure to detect apoptotic smooth and the unknown origin of striated muscle cells during esophageal myogenesis. To reinvestigate this issue, we analyzed esophagi of 4‐day‐old mice by electron microscopy and a fine‐grained sampling strategy considering that, in perinatal esophagus, the replacement of smooth by striated muscle progresses craniocaudally, while striated myogenesis advances caudocranially. We found numerous (1) apoptotic smooth muscle cells located mainly in a transition zone, where smooth intermingled with developing striated muscle cells, and (2) mesenchymal cells in the smooth muscle portion below the transition zone, which appeared to give rise to striated muscle fibers. Taken together, these results provide further evidence for distinct differentiation pathways of both muscle types during esophagus development. Developmental Dynamics 233:964–982, 2005. © 2005 Wiley‐Liss, Inc.
Pink‐eyed dilution alleles affect negative surface charges of mouse spermatozoa
Authors: Hash D. Coleman , Wolfe H. Glenn 
Published: May 2005

AbstractAbnormal spermiogenesis in sterile pink‐eyed dilution mutants results in spermatozoa with bizarre sperm heads. The spermatozoa of normal mice bind colloidal iron hydroxide (CIH) along the length of the tail, yet the spermatozoa of pink‐eyed sterile mice show a great reduction in ability to bind CIH. This implies a loss of negative surface charges. The group(s) responsible for the charges are sensitive to methylation but resistant to neuraminidase treatment, even after deacetylation with alkaline treatment. The membrane components containing the negatively charged groups may be neuraminidase‐resistant forms of gangliosides.
Maternal effect for genes encoding 6‐phosphogluconate dehydrogenase and glucose‐6‐phosphate dehydrogenase in Drosophila melanogaster
Authors: Gerasimova T. I. , Smirnova S. G. 
Published: May 2005

AbstractWe studied the maternal effect for two enzymes of the pentose cycle, 6‐phosphogluconate dehydrogenase (6PGD) and glucose‐6‐phosphate dehydrogenase (G6PD), using a genetic system based on the interaction of Pgd− and Zw− alleles, which inactivate 6PGD and G6PD, respectively. The presence and formation of the enzymes was investigated in those individuals that had not received the corresponding genes from the mother. We revealed maternal forms of the enzymes, detectable up to the pupal stage. The activities of “maternal” 6PGD and G6PD per individual increased 20‐fold to 30‐fold from the egg stage to the 3rd larval instar even in the absence of normal Pgd and Zw genes. Immunologic studies have shown that the increase in 6PGD activity is due to an accumulation of the maternal form of the enzyme molecules. We revealed a hybrid isozyme resulting from an aggregation of the subunits of isozymes controlled by the genes of the mother and embryo itself. These results indicate that the maternal effect in the case of 6PGD is due to a long‐lived stable mRNA transmitted with the egg cytoplasm and translated during the development of Drosophila melanogaster.
Biochemical and genetic analysis of a mutant with altered alkaline phosphatase activity in Dictyostelium discoideum
Authors: MacLeod Carol L. , Loomis William F. 
Published: May 2005

AbstractAlkaline phosphatase is one of several enzymes that accumulate in a temporally regulated sequence during the development of Dictyostelium discoideum. These enzymes can be used to monitor specific gene expression; moreover, isolation and analysis of mutations in the structural gene(s) can serve to indicate some of the essential steps in programmed synthesis and morphogenesis. A mutation (alpA) which affects the activity and substrate affinity of alkaline phosphatase was isolated in D discoideum using a procedure for screening large numbers of clones. Alkaline phosphatase activity at all stages of vegetative growth and development was altered by the mutation. Several physical properties of the enzyme from growing cells and developed cells were compared and found to be indistinguishable. It is likely that a single enzyme is responsible for the majority of alkaline phosphatase activity in growth and development. The mutation is coexpressed in diploids heterozygous for alpA and maps to linkage group III. One of the haploid segregants isolated from these diploids carries convenient markers on each of the six defined linkage groups and can be used for linkage analysis of other genetic loci.
Induction of multiple flagella in Naegleria: Requirements for RNA and protein synthesis
Authors: Walsh Charles , Mar Janet , Ugen Kenneth 
Published: May 2005

AbstractThe induction of multiple flagella by a heat shock was used to examine the role of RNA and protein synthesis in the regulation of the number of flagella produced during the differentiation of the amebo‐flagellate Naegleria gruberi. Control cells differentiating at 25.0°C produce an average of 2.1 flagella per flagellate (f/F), whereas cells exposed to 38.2°C from 35 minutes until 72 minutes after initiation produce an average of approximately 5 f/F. Heat shock was found to prevent completion of the RNA synthesis essential for flagellum formation, even though both RNA and protein synthesis continued at 38.2°C. A delay of two minutes for every one minute of heat shock was seen in both the formation of flagella (T50) and the completion of essential RNA synthesis for heat shocks of one to ten minutes applied beginning 35 minutes after initiation. Times at 38.2°C of ten minutes to 45 minutes produced a delay of approximately 0.6 minutes for each minute of heat shock, whereas shocks longer than 47 minutes prevented flagellum formation. The times from completion of RNA synthesis until completion of protein synthesis or flagella formation were found to be approximately 15 minutes and approximately 32 minutes, respectively. This was true in control cells as well as in cells heat shocked for up to 45 minutes. The fact that heat shock delayed completion of RNA synthesis without affecting the time for completion of protein synthesis or flagella formation suggests that heat shock acts at some step related to the completion of transcription. Short heat shocks, 30 seconds to five minutes, were observed to prevent flagellum formation in cells that had completed RNA synthesis if additional RNA synthesis was inhibited. In contrast, short heat shocks had little effect on cells that had completed protein synthesis, even if additional protein synthesis was inhibited. Two alternative hypotheses for the mechanism of heat shock delay of transcription are discussed. One hypothesis involves a direct effect of high temperature on transcription, and the other postulates a temperature‐sensitive protein necessary for flagellum formation.
Rates of synthesis and turnover of 5′ cap structures of hnRNA and mRNA and their changes during sea urchin development
Authors: Nemer Martin , Ginzburg Irith , Surrey Saul , Litwin Samuel 
Published: May 2005

AbstractThe relationship between heterogeneous nuclear RNA (hnRNA) and messenger RNA (mRNA) synthesis has been studied as a function of the development of the sea urchin embryo through the use of methyl incorporation. Several parameters in the metabolism of capped hnRNA and mRNA of early blastula and late gastrula stages have been investigated by measuring the kinetics of transfer of methyl groups from S‐adenosylmethionine to the 5′ cap structures in nuclear and cytoplasmic RNA:

The rate constants for the decay of hnRNA caps and the synthesis of mRNA caps are equal to within experimental error. This equality indicates a flux of precursor hnRNA caps to mRNA caps with a very high degree of conservation of the hnRNA caps. This conservation holds for each embryonic stage.
From literature data on the labeling kinetics of GTP and mRNA, we have calculated the decay constant of a putative mRNA precursor component of hnRNA. The value of this constant is very close to that for the decay constant of hnRNA caps. Hence, all hnRNA caps and some portion of their associated hnRNA sequences behave kinetically as the pre‐mRNA fraction. This kinetically ascribed pre‐mRNA comprises approximately 30% of the hnRNA mass.
The part of the hnRNA which does not serve as precursor to mRNA turns over at least twice as rapidly as the pre‐mRNA fraction.
During development from early blastula to late gastrula, the rate of hnRNA cap synthesis drops from 2 × 103 molecules/min/cell to half of this value. This decline is parallel to the decline in total hnRNA synthesis and thereby confirms the constant degree of capping of hnRNA, as previously reported. We infer that the pre‐mRNA fraction of hnRNA remains nearly constant during this developmental period.
Specific protection from phenocopy induction by heat shock
Authors: Mitchell Herschel K. , Moller Galina , Petersen Nancy S. , Lipps‐Sarmiento Loveriza 
Published: May 2005

AbstractMild heat treatments applied to whole animals or cell cultures of Drosophila prior to lethal heat shocks result in increased survival and protection against phenocopy induction. The optimal condition for the preliminary mild heat treatment is that which induces the synthesis of heat‐shock proteins but does not turn off the protein synthesis that is in progress. Recovery of protein synthesis but not RNA synthesis following a drastic heat shock is much enhanced by the pretreatments. The results suggest that the protection for survival and against phenocopy induction is due to storage of messenger RNA.
Correlation of phenotypes of the apterous mutation in Drosophila melanogaster
Authors: Wilson Thomas G. 
Published: May 2005

AbstractThe apterous (ap) mutant in Drosophila melanogaster exhibits phenotypes of wing deficiency, precocious adult death, and nonvitellogenic oocyte development. The latter phenotype previously has been shown to result from juvenile hormone (JH) deficiency in the adult stage. To explore the relationship between the hormone deficiency and the other phenotypes, the expression of each phenotype was measured in five alleles of ap (including a new, chemically‐induced allele, ap77f) as wing length, survival five days after eclosion, and initiation and progress of vitellogenic oocyte development. No correlation could be found between severity of wing phenotype and that of precocious adult death or nonvitellogenesis. However, the latter phenotypes were correlated in both ap homozygotes and allelic heterozygotes, since adults that survive have wild‐type vitellogenesis, and those fated for precocious death fail to develop vitellogenic oocytes. These results indicate that no relationship exists between wing and JH deficiencies, but that precocious adult death is related to hormone deficiency — probably through pleiotropy, rather than through causality.
Genetic rescue of lethal genotypes in the mouse
Authors: Gluecksohn‐Waelsch Salome , Teicher Luz S. , Pick Leslie , Cori Carl F. 
Published: May 2005

AbstractDeletions of gene sequences in chromosome 7 of the mouse are known to interfere with biochemical and cellular development differentiation with lethal effects in homozygotes. The presence of the corresponding wild‐type alleles in Cattanach's translocation (chromosomes 7 to X) is able to “rescue” potentially lethal females if they are made heterozygous for the translocation‐carrying X chromosome. This holds true for those chromosome 7 deletions with perinatally lethal effects, whereas “rescue” is not readily accomplished with the deletions that cause early embryonic lethality. Females homozygous for the relevant deletion sequences and heterozygous for the translocation‐carrying X chromosome are mosaics of two cell types: those in which the wild‐type alleles included in the translocated piece complement the depleted sequences, resulting in a normal cellular phenotype, and those with the ordinary X chromosome expressing the lethal phenotype. The developmental interactions between the two cell types and their role in the mechanisms responsible for survival of females homozygous for lethal deletions are discussed. The failure of “rescue” of embryonic lethals reflects as yet unknown temporal and functional aspects of X‐inactivation early embryogenesis.
A β‐glucosidase gene of Dictyostelium discoideum
Authors: Loomis William F. 
Published: May 2005

AbstractSeven mutations affecting β‐glucosidase activity in Dictyostelium discoideum were found to be non‐complementing, recessive to the wild‐type allele, and to occur in the gene locus, gluA. This gene, which is likely to be the structural gene for β‐glucosidase, since a mutation in it gives rise to thermolabile activity and other mutations in it result in no measurable activity, was mapped to linkage group VI. The expression of the β‐glucosidase gene is regulated such that the enzyme is synthesized during the growth phase and during culmination, but not during the first 18 hours following the initiation of development. If expression of the structural gene required the function of a positive regulatory protein coded for by a gene as mutable as the gluA gene, there was greater than 99% chance one of the mutations of this series would have affected the regulatory locus. The absence of a second complementing locus for β‐glucosidase suggests that this enzyme is regulated by other means.
The chorion defect of the ocelliless mutation caused by abnormal follicle cell function
Authors: Underwood E. M. , Mahowald A. P. 
Published: May 2005

AbstractHomozygous Drosophila females bearing the ocelliless mutation are sterile and produce oocytes with abnormal chorions. It has been possible to determine in which tissues these defects reside by generating ovarian chimeras. Pole cells from ocelliless female embryos can give rise to functional oocytes surrounded by normal chorions when placed in a wild‐type environment. Conversely, when wild‐type pole cells are placed in homozygous ocelliless females, the oocytes that form from them have abnormal chorions and never give rise to progeny. Thus the chorion defect and sterility of the ocelliless mutation are not germ‐line autonomous. Homozygous ocelliless ovaries will attach to the uterus when placed in a wild‐type third instar larva, but few eggs are ever laid, and the chorions of stage 14 oocytes remain ocelliless in morphology. Wild‐type ovaries continue to produce oocytes with normal chorion morphology when placed into ocelliless hosts, indicating that the ocelliless chorion defect is ovary autonomous. Thus the chorion defect of the ocelliless mutation resides in the ovarian somatic tissue, presumably the follicle cells.
Evaluation of a catalase inhibitor in the developmental regulation of maize catalase
Authors: Tsaftaris Athanasios S. , Sorenson John C. 
Published: May 2005

AbstractA protein which has been shown to inhibit catalase in vitro appears to vary inversely with catalase activity in the maize scutellum during early sporophytic development when assayed using a catalase inhibition assay. This result suggested that the inhibitor protein may play a direct role in regulating catalase activity during this time period.Four experimental approaches were used to evaluate this putative regulatory role, including immunological quantitation of individual catalase isozymes during germination using rocket immunoelectrophoresis, perturbation of normal catalase expression with hydrogen peroxide or allylisopropylacetamide (AIA), examination of a mutant line with an altered catalase developmental program, and direct radioimmunoassay of the inhibitor protein during germination. The results of these experiments indicate that the quantitative changes in catalase activity during development are not mainly due to changes in the expression of the catalase inhibitor. Other possible roles of this protein in catalase regulation are discussed.
The developmental genetics of pigment mutants in the Mexican axolotl
Authors: Frost Sally K. , Malacinski George M. 
Published: May 2005

AbstractThe Mexican axolotl (Ambystoma mexicanum) provides a well‐defined set of color genes which are useful for various types of analyses. These include the a (albino), m (melanoid), ax (axanthic), and d (white) genes. In addition, various combinations of these genes and a number of as yet undescribed mutants also exist. Three of these mutants (a, ax, and m) have defects associated with specific neural‐crest‐derived pigment cell types. The fourth mutant (d) appears to provide an unsuitable environment for the migration and maintenance of pigment cells. In one case (m), detailed information concerning the specific nature of the genetic defect is available.The goal of this article is to demonstrate ways in which the existing information on the axolotl color genes can best be utilized in terms of understanding not only the mutant phenotypes, but basic concepts in the cell and developmental biology of pigmentation as well. Thus, an attempt has been made to sort through the genetic and biochemical data relevant to these mutants in order to stimulate renewed interest in a more detailed pursuit of such studies.
Role of pupal‐esterase in the regulation of the hormonal status in two Drosophila virilis stocks differing in response to high temperature
Authors: Rauschenbach I. Y. , Lukashina N. S. , Korochkin L. I. 
Published: May 2005

AbstractDrosophila virilis stocks differing in reaction to high temperature (32°C) were studied. The sizes of the larval salivary glands, ring gland, and imaginal discs of the heat‐sensitive stock 147, whose pupal (p) esterase was activated at 32°C, were found to be significantly smaller at high temperature than at 25°C. In larvae of the heat‐resistant stock 101, whose p‐esterase was inactivated at 32°C, the salivary glands and imaginal discs were larger under conditions of high temperature than those of the control larvae. Treatment of stock 147 larvae with ecdysone at 32°C did not affect p‐esterase activity and was 100% lethal. By contrast, the juvenile hormone activated p‐esterase under these conditions and normalized the development of stock 147 larvae. A scheme is suggested for the role of p‐esterase in the regulation of the hormonal status of D. virilis under high temperature conditions.
Control of in vitro growth of viviparous embryo mutants of maize by abscisic acid
Authors: Robichaud C. S. , Wong J. , Sussex I. M. 
Published: May 2005

AbstractThe plant hormone abscisic acid (ABA) is believed to play a role in the onset of developmental arrest in seeds. Embryos of the viviparous mutants of Zea mays do not undergo arrest but germinate directly on the ear. This study investigates the possibility that the mutants vp1, vp5, vp7, vp8, and vp9 are defective in some aspect of ABA action. Mutant and wild type embryos were removed from developing seeds at 18, 21, and 24 days after pollination and cultured aseptically on media containing a range of ABA concentrations. Seedlings were harvested after seven days when lengths and fresh and dry weights were recorded. The results indicate that these five viviparous mutants differ in their response to ABA. Two mutants, vp5 and vp8, exhibit the same sensitivity to growth inhibition by ABA as wild type. The remaining three mutants, however, manifest a range of decreased sensitivities with vp1 being the least sensitive, followed by vp7 and vp9.