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Biomedical subjects

J E Butler

Publications and source records attributed to J E Butler.

At least 91 records · Page 5Linked to original sources
PubMed1994

Characterization of a C alpha gene of swine.

The cDNA sequence encoding the constant region of the porcine IgA heavy chain as well as the exon-intron structure of the germline gene, have been determined. A cDNA clone (1A1) spanning the CH3 domain and part of the CH2 domain was isolated from a porcine mesenteric lymph node cDNA library. Clone 1A1 was aligned with a PCR-generated DNA fragment encompassing the CH1 domain through the 5' end of the CH3 domain to derive the complete cDNA sequence. Comparison with other mammalian C alpha heavy chains (hinge regions excluded) indicated that the deduced amino acid sequence of porcine C alpha is most homologous with the human C alpha subclasses (> 70%), followed by mouse C alpha (61%) and a consensus sequence of the 13 rabbit C alpha heavy chains (59%). The greatest sequence homology was found among the CH3 domains in all species. A striking feature of porcine C alpha is its short six amino acid hinge which like other mammalian IgAs, is encoded with the CH2 domain. Sequence analysis of germline C alpha, generated by PCR from liver or sperm DNA, revealed an exon-intron organization similar to other mammalian C alpha genes. Genomic Southern blot data are consistent with the presence of a single C alpha gene within the porcine genome. Data obtained in these studies will be valuable in pursuing the use of swine as a model in immunological research.

Amino Acid Sequence↗
PubMed1994

Heterogeneity of bovine IgG2. VI. Comparative specificity of monoclonal and polyclonal capture antibodies for IgG2a (A1) and IgG2a (A2).

The relative specificity of 26 randomly selected polyclonal and monoclonal anti-bovine IgG2 reagents for the A1 and A2 allotypic variants of IgG2a was evaluated in a direct RIA using the reagents as solid-phase capture antibodies (CAbs). More than 70% of these reagents were significantly allotype-biased and > 80% of those were positively biased to IgG2a (A1). Compared as the ratio of the ng of IgG2a (A1) bound versus ng IgG2a (A2) bound per 50 ng added (Krel), bias for IgG2a (A1) of six of these reagents was greater than two-fold. Compared in terms of their solid-phase equilibrium constants (Keq), differences as great as two-logs among these reagents were observed. Steward-Petty plots suggested that differences in Krel of a select panel of reagents was usually due to differences in Keq, but for two reagents with large differences in Krel, the existence of one population of CAbs recognizing an allotope and another recognizing common IgG2a determinants, was indicated. Eight of ten guinea pigs immunized with IgG2a (A1) responded with highly significant specificity bias for A1 whereas only two of 11 rabbits and two of ten guinea pigs immunized with IgG2a (A2) responded weakly with preference for IgG2a (A2). These results concur with the concept of the immunodominant nature of the A1 allotope, but also suggest that immunization with IgG2a (A2) might be a practical means of avoiding allotype bias in IgG2a reagents. The data indicate that the majority of randomly selected anti-bovine IgG2 reagents are allotype biased to the extent that when used as serological reagents to measure total IgG2 or bovine IgG2 antibody responses, the allotype of the animal tested rather than its total IgG2a concentration or IgG2 antibody titer, can determine the outcome of the serological test.

Animals↗
PubMed1994

The immunoglobulins and immunoglobulin genes of swine.

The historical works describing the characterization of swine immunoglobulins are reviewed. The three major isotypes, IgM, IgA and IgG, have been recognized for 25 years and their concentrations in various body fluids, the location of the plasma cells throughout the body which synthesize them and their transport into lacteal secretions and absorption by the gut of the newborn piglet, have been studied by many investigators. Swine like humans, have both kappa and lambda light chains and their frequency of expression is similar to that of humans. Various investigators have provided immunochemical evidence for IgG subclass and allotype diversity, although until the recent advent of molecular biology, the complete sequence of any swine immunoglobulin was unknown. Molecular genetic studies reveal single copies of C alpha and C epsilon but as many as eight copies of C gamma. The sequences of five IgG subclasses, IgG1, IgG2a, IgG2b, IgG3 and IgG4, are now available as well as the sequence and genomic organization of C alpha and the sequence of C mu. Swine CH genes all appear to belong to a single small family very similar to human VHIII. Especially interesting is the high degree of similarity among human and swine Ig genes despite the distinct phylogenetic relationship of these species. The rapid expansion of knowledge and technology in the field of molecular biology, together with the attractiveness of the swine as a model for immunoontogeny, in which the influences of both maternal regulatory factors and intestinal gut flora can be experimentally controlled, promises the beginning of an exciting area in swine immunology.

Animals↗
PubMed1993

The immunochemistry of sandwich ELISAs--VI. Greater than 90% of monoclonal and 75% of polyclonal anti-fluorescyl capture antibodies (CAbs) are denatured by passive adsorption.

Quantitative data are presented showing that the method most commonly used to immobilize antibodies in microtiter immunoassays functionally inactivates most of the antibodies. These results were collected using five affinity purified polyclonal antibodies (pAbs) and six monoclonal antibodies (mAbs) specific for fluorescein (FLU) as capture antibodies (CAbs). These CAbs were tested for their ability to capture FLU4.2-BSA after immobilization by passive adsorption, the Protein-Avidin-Biotin-Capture (PABC) system or using previously adsorbed anti-globulins. Results indicate that under optimal conditions, < 10% of monoclonal capture antibody equivalents (CAbeqv) and congruent to 22% of polyclonal CAbeqv remain functional after passive adsorption. Immobilization via the PABC system improved the performance of mAbs by more than five-fold but had less than a two-fold effect on pAbs. Many CAbs immobilized using an anti-globulin retained full activity including the ability to bind two molecules of FLU4.2-BSA/molecule of CAb. The latter result is not necessarily a recommendation for the use of anti-globulin immobilization, since the number of functional CAbeqv per well is not significantly greater than that which can be achieved using passive adsorption.

Adsorption↗
PubMed1992

The physical and functional behavior of capture antibodies adsorbed on polystyrene.

Six monoclonal and two polyclonal antibodies to fluorescein (FLU) were affinity purified and immobilized on Immulon 2 polystyrene as capture antibodies (CAbs): (a) by passive adsorption at pH 9.6, (b) via a streptavidin bridge to a biotinylated carrier molecule, and (c) via an antiglobulin which had been previously adsorbed passively to the polystyrene. Data show that less than 3.0% of the binding sites of monoclonal CAbs and approximately 5-10% of those of polyclonal CAbs were capable of capturing antigen (FLU4.2-BSA) after passive adsorption. Immobilization of CAbs via an antiglobulin or a streptavidin bridge, resulted in the preservation of antibody binding sites to greater than 70% for some monoclonals although immobilization via the streptavidin bridge resulted in the highest number of functional sites/well. The data presented are consistent with studies on other adsorbed proteins which demonstrate that passive adsorption on polystyrene results in the loss of protein function. Furthermore, these data show that generally less than half of the binding sites of antibodies available in solution are available after solid-phase immobilization even when non-adsorptive methods are employed. Some polyclonal anti-FLU also have lower average avidity following passive adsorption compared with CAbs immobilization via a streptavidin bridge. Immunochemical studies revealed that adsorbed polyclonal-CAbs performed like monoclonals when tested with multivalent antigens (FLU10-IgA) but in an expected heterogeneous manner in Scatchard plots when tested using univalent FLU-insulin. This observation implied cross-linking of immobilized CAbs by the multivalent antigen. Because only 5-10% of adsorbed polyclonal CAbs are active, the survivors must be non-randomly distributed in clusters to explain the cross-linking. This was confirmed by scanning electron microscopy which gave rise to the hypothesis that antibodies which retain activity after adsorption, are those present in clusters, i.e., the functional adsorbed CAb is an antibody cluster. Data presented in this report on the behavior of adsorbed CAbs, and reviewed from the work of others for various adsorbed proteins, indicate that the method of passive adsorption at pH 9.6, which is widely used in popular microtiter ELISAs, and which has in many ways revolutionized immunoassay, is a method of protein denaturation. Assayists that utilize passive adsorption of proteins on hydrophobic supports as part of their research need to be cognizant of this phenomenon, while inventors of immunoassay should develop alternative methods of immobilization which do not destroy 90% of the functional activity of solid-phase reactant.

Adsorption↗
PubMed1992

The heterogeneity of bovine IgG2--V. Differences in the primary structure of bovine IgG2 allotypes.

The partial amino acid sequences of the gamma chains of the bovine IgG2a(A1) and IgG2a(A2) allotypes were determined. Sequence differences were found in the CH1 domain, the hinge region, and the CH3 domain. The hinge regions displayed only 71.4% similarity and all of the differences were of a radical nature. The A2 hinge has isoleucine instead of serine at 229, histidine for asparagine at 235, proline for histidine at 238, and cysteine instead of proline in position 234; the latter has the potential for forming an additional interheavy chain disulphide bridge. The occurrence of such a bridge could explain the presence of a pepsin fragment consisting of the hinge region and the Fc. A corresponding fragment is not obtained with the A1 allotype. Both allotypes have a shortened hinge region and a truncated CH2 domain. This feature is characteristic of all reported sequences of IgG2 proteins but not IgG1 in cattle and the goat. This structural feature may be important in subclass-specific recognition by Fc gamma receptors in ruminants. A surprising discovery was the occurrence of five substitutions in the CH3 domain of the IgG2a(A2) in comparison with the A1, which are shared with the CH3 of IgG1. These permit the occurrence of isoallotypic determinants and can explain the difficulty encountered in preparing A2-specific antisera during which adsorption with IgG1 is a routine procedure. The primary sequence data we report confirm the presence of major structural differences between the A allotypes of cattle that was suggested by previous work. The sequence of the A1 allotype most closely agrees with the two IgG2 sequences deduced from their nucleotide sequences whereas the sequence differences in the hinge and C-terminal CH3 make IgG2a(A2) unique. The structural differences between allotypes could have major consequences for such biological activities as phagocytosis, transepithelial transport, lymphocyte and complement activation.

Amino Acid Sequence↗
PubMed1992

Changes in concentration of adenosine triphosphate and adenosine diphosphate in individual preimplantation sheep embryos.

Concentrations of ATP and ADP were measured in 156 sheep embryos by means of an ultramicrofluorescence assay. Stages of preimplantation development measured included unfertilized oocytes through blastocyst-stage embryos. ATP concentrations remained constant through the 8-cell stage; then ATP decreased significantly (p < 0.025) at the morula stage and remained low through the blastocyst stage. ADP concentrations did not change throughout the embryonic stages measured. Decreased levels of ATP with constant levels of ADP caused the ATP:ADP ratio to decrease significantly (p < 0.025) between the 8-cell and morula stages. We suggest that the increase in glucose uptake by sheep embryos observed at the morula stage of development may be due, in part, to a decrease in the ATP:ADP ratio.

Adenosine Diphosphate↗
PubMed1991

Growth and DNA replication in rabbit blastocysts.

DNA content and DNA polymerase activity were measured on rabbit blastocysts removed from the uterus at 24-hr intervals over the period of days 4-7 postcoitum (pc). Median DNA content increased 53 times over the 72-hr period, from 25.3 ng on day 4 to 1,360 ng on day 7. Median DNA polymerase activity (fmole of radiolabeled nucleotide incorporated in 30 min at 37 degrees C) increased 393-fold from day 4 to day 7: 32.8 to 12,900. These embryos also increased in surface area and volume by 334-fold and 6,078-fold, respectively. Litters containing individuals with high DNA content also tended to have similar individuals with high DNA polymerase activity. Therefore, DNA polymerase activity may be a useful measure of the potential for the next cell division. A large amount of variation existed between blastocysts in all parameters measured. An analysis of variance, conducted to partition variation between litters and within litters, determined that within-litter variation was actually greater than that between litters, resulting in intraclass correlation coefficients less than 0.5. There was also a positive regression of DNA content and DNA polymerase activity on surface area in 6- and 7-day-old blastocysts after eliminating variation attributable to litters. The developmental pattern of DNA polymerase activity in the rabbit may be quantitatively different from that described in the mouse. The pattern in mammals is very different from that described in several nonmammalian species.

Animals↗
PubMed1991

Co-culture of day-5 to day-7 equine embryos in medium with oviductal tissue.

Oviductal and uterine embryos were collected from mares at 5 to 7 days following ovulation 1) to evaluate the effects of oviductal tissue explants on in vitro growth and development of equine embryos and 2) to study the morphologic development of equine embryos in culture. Embryos were incubated for 5 days in a medium (control group) or in medium supplemented with oviductal tissue explants (co-culture group). Embryos were evaluated and the media changed daily. Following 5 days in culture, 10 10 (100%) control embryos and 27 29 (93%) co-cultured embryos had doubled in diameter. All embryos that were recovered as morulae developed to the blastocyst stage in culture. By 5 days in culture, 6 10 (60%) control embryos and 19 29 (66%) co-cultured embryos had reached the hatching blastocyst stage of development. By 3 days in culture, significantly more (P<0.05) control embryos versus co-cultured embryos had degenerated (4 10 vs 2 29 , respectively). By 5 days in culture, significantly more (P<0.01) control embryos versus co-cultured embryos had degenerated (6 10 vs. 3 29 , respectively). Embryos cultured with oviductal tissue were sustained longer than embryos cultured in medium alone. Hatching was characterized by the blastocyst squeezing through a small opening in the zona pellucida or by the zona pellucida thinning over approximately half of the blastocyst surface and subsequently disappearing entirely.

Journal Article↗
PubMed1990

Preliminary characterization of pyruvate uptake by one-cell ovine embryos.

Pyruvate uptake was measured by a noninvasive method on a total of 59 individual one-cell ovine embryos. Uptake was measured over a 4-h period in Whitten's medium containing 1.0 mM glucose and either 0.0 mM, 0.33 mM, 0.67 mM or 1.0 mM sodium pyruvate. Glucose uptake was also assessed to determine the effect of altering pyruvate concentration on glucose uptake. Embryos incubated in medium containing 1.0 mM or 0.67 mM pyruvate showed average uptake values of 20.36 and 5.00 pmoles per embryo per 4 h, respectively. In contrast, when pyruvate concentrations were 0.0 mM and 0.33 mM, the embryos released an average of 24.60 and 10.40 pmoles pyruvate per embryo per 4 h. Uptake or release of pyruvate was found to vary linearly with concentration level. Increasing pyruvate concentration resulted in a significant decrease in glucose uptake.

Journal Article↗
PubMed1990

The antigen-limited nature of microtiter ELISAs requires partial depletion of IgG to permit reliable determination of rabbit serum IgA antibody activity.

The antigen-limiting nature of microtiter ELISAs predicts that antibodies of minor classes may be underestimated when the same specimen contains large amounts of IgG antibodies specific for the same antigen. Such competitive inhibition can be diagnosed from ELISA titration plots. A method is described to eliminate the negative effects of this competition on the detection of IgA antibodies in rabbit serum. The detectability of rabbit serum antibodies to ovalbumin and bovine serum albumin is increased 10-fold by prior treatment of 1:100 dilutions of serum with 1% Cowan I S. aureus. High concns of S. aureus, e.g. 10%, completely deplete serum IgG without loss of IgA. However, concns higher than 1% do not lead to additional improvement in the detectability of IgA antibodies in the systems studied. The method is rapid, inexpensive and shows no non-specific depletion of IgA from either serum or bronchoalveolar lavage fluid.

Animals↗
PubMed1990

Fluoride: an adjuvant for mucosal and systemic immunity.

Fluoride, the agent responsible for reduction of dental caries worldwide, and a recognized proliferative agent, is a potent adjuvant when given intragastrically to rats. Intragastric fluoride causes increases in the size and cellularity of the Peyer's patches and mesenteric lymph nodes as well as the number of plasma cells secreting IgG and IgA antibodies to ovalbumin given in their drinking water. Rats ingesting NaF and fed OA showed a significant increase in surface immunoglobulin expression on lymphocytes from the Peyer's patches and mesenteric lymph nodes. The frequency of CD4+ T cells in these lymphoid tissues was elevated while that of CD8+ T cells was significantly decreased. In separate experiments, rats parenterally immunized with myelin basic protein (MBP) and fed NaF twice weekly, had significantly elevated serum IgG antibody activity to MBP compared to similarly immunized rats not receiving NaF. The supplemental fluoride prescribed for infants and especially that which is inadvertently ingested by children and adults given fluoride gels, is within the concentration range of that which produced the effects we observed in rats. The adjuvant effect we describe thus has relevance for fluoride therapy worldwide.

Adjuvants, Immunologic↗
PubMed1990

Humoral immunity in root caries in an elderly population. 1.

IgA, IgG and IgM antibody activity (ELISA Units/ml) to Streptococcus mutans, Actinomyces viscous and Escherichia coli CF8 in serum, parotid saliva and whole saliva was measured using the amplified ELISA (a-ELISA) while the concentration (microgram/ml) of each isotype of immunoglobulin as well as albumin and lactoferrin, was determined using sandwich ELISAs. Selection of suitable reagents from those commercially available was based on specificity tests using purified human immunoglobulin; most polyclonal reagents required further absorption to attain class specificity. Cross-absorption studies indicated the absence of patient antibodies that were cross-reactive among the bacteria studied, except for IgM in some cases. Expression of response in ELISA Units (E.U.) per microgram of immunoglobulin, i.e. specific activity, revealed that IgG specific activity was significantly higher in parotid saliva than in either whole saliva or serum for all bacteria studied; serum and whole saliva did not differ except for the higher specific activity in whole saliva to E. coli. The value of one E.U. was determined using the Comparative Antibody-immunoglobulin Capture Assay (CACA). Using this novel method, we estimated that about 0.05 percent of serum IgA was specific for Streptococcus mutans, 0.008 for Actinomyces viscosus and 0.004 for Escherichia coli CF8. The percentage of specific IgM antibodies was higher than for IgA and IgG. The concentration of IgA anti-Streptococcus mutans, Actinomyces viscosus and Escherichia coli levels are approximately 92 ng/ml, 25 ng/ml and 16 ng/ml in whole saliva and 46 ng/ml, 9.4 ng/ml and 6.3 ng/ml in parotid saliva.(ABSTRACT TRUNCATED AT 250 WORDS)

Actinomyces↗
PubMed1990

Humoral immunity in root caries in an elderly population. 2.

Saliva specimens stored for 18 months at -20 degrees C with or without glycerol and the anti-protease benzamidine-HCl, lost all antibody activity for S. mutans. IgA activity in processed whole saliva decreased significantly after one week when stored either at 4 degrees C or -20 degrees C with or without glycerol, although it was stable in parotid saliva for at least 40 days. Loss of activity prior to processing was significant in the first 24 h, and the addition of 50% glycerol and storage at -70 degrees after processing, prevented loss of antibody activity in both whole and parotid saliva. Diurnal variations in IgA, lactoferrin and the IgA secretion rate were insignificant in parotid saliva but showed some fluctuations in whole saliva. Albumin and lactoferrin levels exhibited the greatest fluctuation in whole saliva specimens although IgA and IgA antibody levels were still more characteristic of the patient than the time of sampling. Monthly variations in IgA, IgA antibody activity and other parameters were least in parotid saliva and e.g., values for parameters that were high in patients samples on the first month, remained high during the 4-month study period. Statistical analyses showed a high correlation between values obtained for most of the 15 parameters that were measured in parotid and whole saliva specimens collected from greater than 20 patients during 2 successive visits. Whole saliva values for albumin, lactoferrin and albumin levels in parotid saliva, were most variable but differences were not significant. Hence, patients with very low or very high values, even in whole saliva, can be identified within the population on the basis of specimens collected at a single time.

Aged↗
PubMed1990

Maternal-neonatal immunoregulation: suppression of de novo synthesis of IgG and IgA, but not IgM, in neonatal pigs by bovine colostrum, is lost upon storage.

Fifty-four neonatal pigs were allotted to 4 groups and reared in an electrically controlled automatic feeding device (autosow). Each group was reared on a different pool of bovine colostrum: fresh, stored 1 month, stored 6 months, and stored 8 years. Bovine and porcine immunoglobulins in the sera of these pigs, and in a group of conventionally reared pigs, were measured periodically during the first 42 days after birth. The maximal concentration of absorbed bovine immunoglobulin was reached between 12 and 18 hours and equaled or exceeded the amount of porcine immunoglobulin absorbed by the conventionally reared pigs. Large differences in the concentrations of the bovine immunoglobulin isotypes among the various pools of colostrum were positively correlated with concentration of these isotypes in the sera of the neonatal pigs fed these pools. Relative to their concentrations in colostrum, approximately 41% of the IgG1, 55% of the IgG2, 29% of the IgM, and 67% of the IgA was absorbed. The IgA was absorbed the best and IgM was least absorbed. Significant trends or differences in absorption were not observed among groups. Neonatal pigs given fresh colostrum, which had a higher fat content, had significantly more weight gain (P less than 0.05). This occurred, despite the fact that the fresh colostrum had the lowest concentration of bovine immunoglobulin. Serum half-lives for bovine IgG1 and IgG2 were significantly less than for porcine IgG (P less than 0.05), whereas the half-lives for bovine and porcine IgM and IgA were similar. De novo-synthesized immunoglobulins were detectable in serum after 6 days; IgM concentrations reached a maximum at 15 days in neonatal pigs given stored, but not fresh, colostrum.(ABSTRACT TRUNCATED AT 250 WORDS)

Animals↗
PubMed1989

The immunochemistry of sandwich ELISAs--III. The stoichiometry and efficacy of the protein-avidin-biotin capture (PABC) system.

The protein-avidin-biotin capture (PABC) system was developed to decrease the adsorption-induced loss of antigen capture capacity (AgCC) of capture antibodies (CAb) used in sandwich ELISAs. This system involves immobilization of biotinylated CAbs through linkage by streptavidin (SA) to biotinylated carrier proteins adsorbed on polystyrene. Studies reported here describe the stoichiometry of the system and the influence of biotinylation of different carrier proteins and CAbs on the reaction stoichiometry and the AgCC of CAbs. Because of the widespread use of sandwich ELISAs to measure the concn of multivalent protein antigens, the AgCCs of monoclonal and polyclonal CAbs to pig IgG in the PABC system were compared with the AgCCs of these Abs immobilized on the plastic by direct adsorption. Optimal assay conditions for the carrier were obtained when 1 microgram/ml of the biotinylated protein was added to the polystyrene solid phase. An increasing degree of biotin substitution in three carrier proteins was paralleled by an increasing AgCC until a constant maximum was reached. Under conditions of maximal AgCC, 120 ng of the carrier rabbit gamma globulin (RGG; i.e. RGG25biot) was bound to polystyrene, which in turn yielded the maximum amount (i.e. 100 ng) of bound streptavidin (SA; Bdngmax) when 20 micrograms/ml of SA was added. Under conditions giving the Bdngmax for SA, CAb12biot yielded a higher Bdngmax than did CAb25biot or CAb2biot. When the AgCC of equal amounts of differentially biotinylated CAbs were compared, the following order of AgCC was observed: CAb12biot greater than CAb12biot greater than CAb25biot. Hence, while the maximal amount of CAb is immobilized on SA when CAb12biot is used, optimal AgCC is achieved with CAb2biot. The carrier:SA:CAb2biot ratio was 1:2:1 while that for carrier:SA:CAb12biot was 1:2:2. The same ratio was obtained using IgG2biot from four different species. Monoclonal antibodies to swine IgG showed a 5-6-fold increase in Bd%max when immobilized as CAbs using the PABC system versus when adsorbed on polystyrene. Plots of these data suggest that the differences result from a loss of functional affinity. On the contrary, no significant differences in Bd%max and hence functional affinity were observed when a polyclonal antibody to pig IgG was compared using the two assay configurations. Furthermore, when the globulin fraction of the anti-pig polyclonal was adsorbed on plastic, it behaved nearly as well as its affinity-purified counterpart immobilized by the PABC system. The PABC system appears to offer significant advantages for sandwich ELISAs utilizing monoclonal antibodies as the CAb, and may offer some advantages in other s

Adsorption↗
PubMed1989

Application of theoretical considerations to the analysis of ELISA data.

Solid-phase immunoassays such as the ELISA are in routine use in many areas of biological research. Data from these assays are analyzed in a variety of ways, frequently without taking into account the immunochemical principles of the assay. The Reference Standard Method is often used and is suitable and convenient for obtaining concentration (or activity) values from the antigen-specific ELISA or spRIA, sandwich assays, and inhibition assays. The standard curve required for this method may be obtained by simple linear regression analysis of logarithmic or logitlogarithmic transformed data obtained from titration of the reference standard. The shape of the logarithmic plot of the reference standard provides information on the performance of the assay. Examining data from multiple dilutions of the samples is essential to assure that each titrates with the same slope as does the reference standard; the analysis routine must permit this comparison to be made. ELISANALYSIS is a program for the IBM PC which was developed to perform such analyses. It is presented here as a model, with sufficient information provided for the development of similar analytical routines by interested users. This approach to ELISA data analysis is presented as an alternative to complicated empirical curve-fitting systems and simple endpoint methods, which can be immunochemically misleading or, in some cases, even invalid. The consistent use of the described routines would encourage greater uniformity in the means of data interpretation and thereby enhance our understanding of immunobiology.

Algorithms↗