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The papers listed below are available for download in Adobe Reader *.pdf format. You may view and print these files with the freely redistributable Reader from Adobe.

  • Taswell C, 1987, Journal of Immunology 138(2):333-341.
    A solution to the problems of cytolysis assays with additional applications to other immunological and biochemical assays.

    After cellular immunoassays are compared with classical bioassays, conventional methods and consequent problems of data analysis for cytolysis assays are reviewed and a new solution is proposed. This solution incorporates new methods, called dose-response surface assays and analysis (DRSA), which estimate cytolytic activity coefficients on a surface in a three-dimensional space with two dose variables (killers and targets) and one response variable (counts). These new methods based on dose-response surfaces are demonstrated to be more informative and reliable than classical methods based on dose-response curves. In a test of the methods' robustness (sensitivity of parameter estimates to changes in the dose levels of the assay design), cytolytic activity coefficients estimated by DRSA varied by ≤30% over a reduction of three to four orders of magnitude in the dose levels. This remarkable robustness should be compared with the corresponding figures of as much as 500% over less than <1 order of magnitude for previously published results of coefficients estimated by conventional methods. DRSA is distinguished from replot-of-plots methods such as those used for enzyme inhibition assays in biochemistry, and is recommended as a more efficient method that should replace replot-of-plot methods now antiquated by the advent of microcomputers. DRSA can be applied to any experimental system that requires an activity coefficient to be estimated on a dose-response surface in a space of ≥3 dimensions (≥2 dose variables and one response variable), regardless of the mathematical model and statistical estimators used to analyze the dose-response interaction. Finally, DRSA is compared with the methods known as response surface methodology (RSM), and is described as a new class of methods to be added to those that constitute RSM.

  • Taswell C, 1987, Cell Separation Methods and Selected Applications 4:109-145.
    Limiting dilution assays for the separation, characterization, and quantitation of biologically active particles and their clonal progeny.

    This review chapter was published without a separate abstract. Excerpts from the introduction and conclusion are available on the LDA Review Page.

  • Taswell C, 1984, Journal of Immunological Methods 72(1):29-40.
    Limiting dilution assays for the determination of immunocompetent cell frequencies. III. Validity tests for the single-hit Poisson model.

    A statistical method was developed to test the validity of the single-hit Poisson model in limiting dilution assays used to determine immunocompetent cell frequencies. Principles of bioassay, validity tests, and the distinction between model-discrimination experiments and parameter-estimation assays are reviewed in the Introduction. The new test derived and then demonstrated with previously published data is intended to be used for parameter-estimation assays based upon the single-hit Poisson model. It is a family of related χ2, t, and F tests for deviations from zero of the slopes of weighted least squares regression plots. These plots regress the logarithms of single-dose estimates fi of the frequency φ on the total cell doses λi and fi on the total cell dose reciprocals 1/λi, that is, Yi = ln fi on Xi = λi and Yi = fi on Xi = 1/λi. The test discriminates against alternative models with multiple-hit/target response-generation processes, a variable number (dose-dependent) of false negatives, and a constant number (dose-independent) of false positives. Its purpose as a test for parameter-estimation assays, though, is to detect deviations from the single-hit Poisson model and not to select one of these alternative models. Tests for model-discrimination experiments to select or 'prove' an unknown alternative model are considered in light of relevant literature reviewed in the Discussion.

  • Taswell C, 1984, M.Sc. Thesis, Department of Mathematics, New York University.
    Limiting dilution assays for the determination of immunocompetent cell frequencies. II. Experimental design for single-dose assays.

    A statistical method was developed to optimize experimental designs for quantal response limiting dilution assays used to determine the frequencies of immunocompetent cells. Formulas for the design optimization criteria were derived by incorporating a prior distribution for the immunocompetent cell frequency into the design optimization analysis. This prior distribution was used to parameterize the advance information known about the frequency. The Cramer-Rao minimum variance and uninformative assay probability were chosen as the optimization criteria. Tables of these parameters, their corresponding optimum doses, and other experimental design parameters were computed for various frequency prior distributions. This method was developed in order to resolve the problems of previous approaches. The values computed for the optimum efficiency dose and for the minimum error of the frequency estimate are respectively higher and lower than those obtained by previous authors. Additional results predict that it is possible to perform single-dose assays successfully (with low uninformative assay probability) at doses approaching the optimum efficiency dose. (Single-dose assays should be used, however, only when the single-hit Poisson model used to estimate the frequency has already been proven valid by a multiple-dose experiment. References are provided for these validity tests.) To optimize the design of future assays in a sequence, information from past assays in the sequence should be used with the method and results presented here. The errors of frequency estimates could then theoretically be reduced to levels lower than those previously considered attainable.

  • Taswell C, 1981, Journal of Immunology 126(4):1614-1619.
    Limiting dilution assays for the determination of immunocompetent cell frequencies. I. Data analysis.

    A statistical method was developed for the analysis of experimental data from limiting dilution assays. Formulas for the estimation of the frequency of immunocompetent cells within a test population were derived by the statistical methods of weighted averaging, likelihood maximization, and χ2 minimization. Equations for the latter 2 were solved by Newton's method of iterative approximation. Estimates obtained by these methods were found to be more valid than those obtained by least squares (LS) fitting as judged by the χ2 test and as established by Monte Carlo experiments. χ2 minimization was chosen as the preferable estimation method with maximum accuracy and precision (minimum bias and variance) for the standard determination of frequencies; likelihood maximization was used only for the confirmation of results. When data from previously published experiments were reanalyzed, both results and conclusions were found to differ significantly from those originally obtained by LS fitting, thus demonstrating the importance of using proper data analysis methods. In conjunction with the use of available calculators or microcomputers, the method presented here provides a simple and rapid procedure for the valid determination of immunocompetent cell frequencies.

  • Taswell C, MacDonald HR, Cerottini JC, 1980, Journal of Experimental Medicine 151(6):1372-1385.
    Clonal analysis of cytolytic T lymphocyte specificity. I. Phenotypically distinct sets of clones as the cellular basis of cross-reactivity to alloantigens.

    The cellular basis of the cytolytic cross-reactivity observed in primary allogeneic (C56BL/6 anti-DBA/2 and C57BL/6 anti-C3H/He) mixed-leukocyte cultures (MLC) was investigated by analysis of the specificity of clonal progeny derived from individual cytolytic T lymphocyte (CTL) precursor cells (CTL-P) contained within these populations. A sensitive mixed-leukocyte microculture (micro-MLC) technique was used with limiting dilution analysis by Poisson statistics to determine the frequency of CTL-P reactive against both specific and third-party (P815 and AKRA) target cells, to calculate the probability that each micro-MLC was a clone derived from a single CTL-P, and to examine the specificity of each micro-MLC assayed separately against both target cells. A total of 287 phenotypically specific, heteroclitic, and cross-reactive micro-MLC from the 2 different strain combinations were observed with a relative frequency of 81, 11, and 8%, respectively, and were calculated to have mean clone probabilities of 90 and 99% when based, respectively, upon the frequencies of CTL-P reactive against the specific and third-party target cells. These clones were estimated to have an approximate size of 6 x 104 cells, which corresponded to roughly 16 cell doublings during the 7 d of culture. 22 clones were successfully subcloned and in virtually every case, the subclones retained the specificity phenotype of the original clone from which they were derived. These results provide direct evidence for three phenotypically distinct sets of CTL as the cellular basis of cross-reactivity in MLC populations assayed against two different target cells.

  • Taswell C, MacDonald HR, Cerottini JC, 1979, Thymus 1(1-2):119-131.
    Limiting dilution analysis of alloantigen-reactive T lymphocytes. II. Effect of cortisone and cyclophosphamide on cytolytic T lymphocyte precursor frequencies in the thymus.

    A minimal estimate of the frequency of cytolytic T lymphocyte precursors (CTL-P) in the thymus was determined by application of Poisson statistics to limiting dilution analysis. A mean CTL-P frequency of 1/1467 was obtained for C57BL/6 (H-2b) thymus cells activated by DBA/2 (H-2d) irradiated spleen cells and assayed against P-815 mastocytoma (H-2d) target cells. CTL-P frequencies were also obtained for spleen, nylon wool column purified spleen, peripheral blood, and lymph node cell populations. The effect of in vivo drug treatments on CTL-P frequencies was then examined. Cortisone at 100 mg/kg dramatically increased the CTL-P frequency in thymus by more than 20-fold despite a drastic reduction in the number of total thymus cells. The same cortisone treatment did not affect the CTL-P frequency in spleen. In contrast, cyclophosphamide at 300 mg/kg decreased the CTL-P frequency in spleen by more than 10-fold without affecting that in thymus. Cyclophosphamide at 100 mg/kg did not produce any significant change. A detailed explanation of the calculation of CTL-P frequencies is provided and their validity is discussed.

  • Taswell C, McDuffie FC, Mann KG, 1975, Immunochemistry 12(4):339-343.
    Immunochemical relationships of the intermediates of prothrombin activation.

    The mechanism of prothrombin activation previously reported from this laboratory (Heldebrant et al., 1973) has been immunochemically verified by analysis of the precursor, intermediate, and product relationships of the proteins with a micro Ouchterlony precipitin test. The cross reactions between purified bovine prothrombin, intermediates of activation, and thrombin were examined with rabbit and chicken antisera to each of these proteins. In each instance, the derivative was found to cross react with its parent protein, yet was found to be completely distinct from any other protein derived from the same cleavage. The results support a mechanism in which prothrombin (70,000 daltons) is cleaved to yield intermediates 1 (51,000 daltons) and 3 (23,000 daltons); intermediate 1 is then cleaved to yield intermediates 2 (41,000 daltons) and 4 (13,000 daltons); thrombin (39,000 daltons) is finally produced from intermediate 2.

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