ABSTRACT
BACKGROUND: DNA analysis of endoreduplicating cells is difficult because of the overlap between stem-line G2 + M cells and 4C G1 cells. Simultaneous flow cytometry of DNA and cyclin B1 analytically separates these populations. The objective here was to develop simultaneous flow cytometry of DNA, cyclin B1, and p105 (highly expressed in mitosis) for improved, complete cell cycle phase fraction analysis of endoreduplicating cell populations. METHODS: Monoclonal antibody, GNS-1, reactive with human cyclin B1, was conjugated with fluorescein at three different fluorochrome-to-protein (F/P) ratios and tested for optimal sensitivity in a flow cytometric assay. A formaldehyde-methanol fixation procedure was optimized for retention of p105 within mitotic cells by analytic titration of formaldehyde. p105 was stained indirectly with Cy5-conjugated secondary antibody, followed by GNS-1, and DNA was stained with Hoechst 33342. The specificity of p105 in this assay was tested by comparison of manual and flow cytometric mitotic indices and by sorting and microscopic inspection. RESULTS: F/P 4.1 provided optimal fluorescein labeling of GNS-1. Formaldehyde (0.5%), followed by methanol permeabilization, fixed cells sufficiently to quantify stem-line and endoreduplicated G1, S, G2, and M phase fractions. Kinetic measurements of these fractions for both populations were demonstrated. CONCLUSIONS: The fluorochrome-to-protein ratio is important and can be optimized objectively for these assays. A permeabilization-sensitive antigen (p105), previously requiring formaldehyde/detergent-fixed cell preparations, was shown to work equally well with formaldehyde/ methanol fixation. Three-laser, two-parameter intracellular antigen analysis can be successfully coupled with DNA content analysis. Cell cycle kinetic analysis of endoreduplicating populations should be improved.
Subject(s)
Cell Cycle/physiology , Cyclin B/analysis , DNA/analysis , Flow Cytometry/methods , Retinoblastoma Protein/analysis , Benzimidazoles/metabolism , Cell Line , Cell Separation , Cells, Immobilized , Cyclin B1 , Dose-Response Relationship, Drug , Fluorescein/metabolism , Fluorescein-5-isothiocyanate/metabolism , Formaldehyde/metabolism , Humans , Immunohistochemistry , Male , Methanol/metabolism , Prostatic Neoplasms/metabolism , Time Factors , Tumor Cells, CulturedABSTRACT
Despite significant improvements in the wear resistance of posterior composite restorations, they still undergo occlusal wear, color change, and surface staining with time. One method of repairing these restorations is re-surfacing the old composite. This study investigated the bonding of new composite to the corresponding old composite material by several different mechanical conditioning steps, chemical conditioning steps, primer conditioning steps, and four posterior composites. Aged composite surfaces were conditioned, re-bonded to new composite, stored in artificial saliva for seven days at 37 degrees C, and tested in shear. The mean shear strength for unrepaired composites was 27 MPa. Optimal re-bond strengths were 88% (Estilux), 77% (Ful-Fil), 92% (Occlusin), and 102% (P-10) of original bulk shear strengths. General linear modeling revealed that the best combination tested was roughening with a D830 diamond bur, conditioning the surface with water, and priming with Scotchbond-1 dentin bonding agent.
Subject(s)
Composite Resins , Dental Bonding/methods , Dental Restoration, Permanent/methodsSubject(s)
Composite Resins , Dental Restoration, Permanent , Resins, Synthetic , Clinical Trials as Topic , Color , Cuspid , Dental Caries , Follow-Up Studies , Incisor , Tooth AbrasionABSTRACT
Acquired immunodeficiency syndrome (AIDS) has become one of the most pressing public health issues of this decade. As a member of the health care team, dental professionals are increasingly confronted with the responsibilities of treating AIDS victims, preventing the transmission of HIV and other infectious agents, and being nondiscriminatory employers. Guidelines are currently available to help the dental profession in these first two responsibilities. However, policies governing the employment of health care workers with AIDS have not been established firmly. Therefore, this article focuses on the AIDS employment issue facing dental practitioners, administrators, and educators today. Background information on the current medical status, transmission, epidemiology, and populations-at-risk is reviewed. Legal considerations of AIDS in the work place are addressed as to HIV testing, employee rights to gain and/or retain employment, economic considerations, and confidentiality issues.
