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1.  Geographic Divergence of Bovine and Human Shiga Toxin–Producing Escherichia coli O157:H7 Genotypes, New Zealand1 
Emerging Infectious Diseases  2014;20(12):1980-1989.
A historically introduced subset of globally circulating strains continue to evolve and be transmitted between cattle and humans.
Shiga toxin-producing Escherichia coli (STEC) O157:H7 is a zoonotic pathogen of public health concern worldwide. To compare the local and large-scale geographic distributions of genotypes of STEC O157:H7 isolates obtained from various bovine and human sources during 2008–2011, we used pulsed-field gel electrophoresis and Shiga toxin–encoding bacteriophage insertion (SBI) typing. Using multivariate methods, we compared isolates from the North and South Islands of New Zealand with isolates from Australia and the United States. The STEC O157:H7 population structure differed substantially between the 2 islands and showed evidence of finer scale spatial structuring, which is consistent with highly localized transmission rather than disseminated foodborne outbreaks. The distribution of SBI types differed markedly among isolates from New Zealand, Australia, and the United States. Our findings also provide evidence for the historic introduction into New Zealand of a subset of globally circulating STEC O157:H7 strains that have continued to evolve and be transmitted locally between cattle and humans.
doi:10.3201/eid2012.140281
PMCID: PMC4257794  PMID: 25568924
molecular epidemiology; Shiga toxin–producing Escherichia coli O157:H7; genotype; Shiga toxin–encoding bacteriophage insertion typing; cattle; transmission; New Zealand; geographic divergence; population structure; proportional similarity index; enteric infections; bacteria
2.  A prospective case–control and molecular epidemiological study of human cases of Shiga toxin-producing Escherichia coli in New Zealand 
BMC Infectious Diseases  2013;13:450.
Background
Shiga toxin-producing Escherichia coli (STEC) O157:H7 and related non-O157 STEC strains are enteric pathogens of public health concern worldwide, causing life-threatening diseases. Cattle are considered the principal hosts and have been shown to be a source of infection for both foodborne and environmental outbreaks in humans. The aims of this study were to investigate risk factors associated with sporadic STEC infections in humans in New Zealand and to provide epidemiological information about the source and exposure pathways.
Methods
During a national prospective case–control study from July 2011 to July 2012, any confirmed case of STEC infection notified to regional public health units, together with a random selection of controls intended to be representative of the national demography, were interviewed for risk factor evaluation. Isolates from each case were genotyped using pulsed-field gel electrophoresis (PFGE) and Shiga toxin-encoding bacteriophage insertion (SBI) typing.
Results
Questionnaire data from 113 eligible cases and 506 controls were analysed using multivariate logistic regression. Statistically significant animal and environmental risk factors for human STEC infections were identified, notably 'Cattle livestock present in meshblock’ (the smallest geographical unit) (odds ratio 1.89, 95% CI 1.04–3.42), 'Contact with animal manure’ (OR 2.09, 95% CI 1.12–3.90), and 'Contact with recreational waters’ (OR 2.95, 95% CI 1.30–6.70). No food-associated risk factors were identified as sources of STEC infection. E. coli O157:H7 caused 100/113 (88.5%) of clinical STEC infections in this study, and 97/100 isolates were available for molecular analysis. PFGE profiles of isolates revealed three distinctive clusters of genotypes, and these were strongly correlated with SBI type. The variable 'Island of residence’ (North or South Island of New Zealand) was significantly associated with PFGE genotype (p = 0.012).
Conclusions
Our findings implicate environmental and animal contact, but not food, as significant exposure pathways for sporadic STEC infections in humans in New Zealand. Risk factors associated with beef and dairy cattle suggest that ruminants are the most important sources of STEC infection. Notably, outbreaks of STEC infections are rare in New Zealand and this further suggests that food is not a significant exposure pathway.
doi:10.1186/1471-2334-13-450
PMCID: PMC3854066  PMID: 24079470
Prospective case–control study; Sporadic STEC infections; New Zealand; Risk factors; Source attribution; Cattle; Molecular epidemiology; Pathways of infection; Population attributable fractions
4.  The “structurally minimal” isoform KChIP2d modulates recovery of Kv4.3 N-terminal deletion mutant Δ2-39 
Channels  2011;5(3):225-227.
Mechanisms underlying Kv4 (Shal type) potassium channel macroscopic (open state) inactivation and recovery are unknown, as are mechanisms by which KChIP2 isoforms modulate these two processes. In a recent study (Xenopus oocytes, 2 microelectrode voltage clamp) we demonstrated that: i) Partial deletion of the Kv4.3 proximal N-terminal domain (Δ2-39; deletes N-terminal amino acids 2–39) not only slowed macroscopic inactivation, but also slowed the net rate of recovery; and ii) Co-expression of KChIP2b significantly accelerated the rate Δ2-39 recovery from inactivation. The latter effect demonstrated that an intact N-terminal domain was not obligatorily required for KChiP2b-mediated modulation of Kv4.3 recovery. To extend these prior observations, we have employed identical protocols to determine effects of KChiP2d on Δ2-39 macroscopic recovery. KChiP2d is a “structurally minimal” isoform (consisting of only the last 70 amino acids of the common C-terminal domain of larger KChIP2 isoforms) that exerts functional modulatory effects on native Kv4.3 channels. We demonstrate that KChiP2d also accelerates Δ2-39 recovery from macroscopic inactivation. Consistent with our prior Δ2-39 + KChIP2b study, these Δ2-39 + KChIP2d results: i) Further indicate that KChIP2 isoform-mediated acceleration of Kv4.3 macroscopic recovery is not obligatorily dependent upon an intact proximal N-terminal; and ii) Suggest that the last 70 amino acids of the common C-terminal of KChiP2 isoforms may contain the domain(s) responsible for modulation of recovery.
doi:10.4161/chan.5.3.15313
PMCID: PMC3225751  PMID: 21422811
Kv4.3; KChIPs; inactivation; recovery; potassium channels
5.  KV4.3 N-terminal deletion mutant Δ2–39 
Channels  2011;5(1):43-55.
Gating transitions in the KV4.3 N-terminal deletion mutant Δ2–39 were characterized in the absence and presence of KChIP2b. We particularly focused on gating characteristics of macroscopic (open state) versus closed state inactivation (CSI) and recovery. In the absence of KChIP2b Δ2–39 did not significantly alter the steady-state activation “a4” relationship or general CSI characteristics, but it did slow the kinetics of deactivation, macroscopic inactivation and macroscopic recovery. Recovery kinetics (for both WT KV4.3 and Δ2–39) were complicated and displayed sigmoidicity, a process which was enhanced by Δ2–39. Deletion of the proximal N-terminal domain therefore appeared to specifically slow mechanisms involved in regulating gating transitions occurring after the channel open state(s) had been reached. In the presence of KChIP2b Δ2–39 recovery kinetics (from both macroscopic and CSI) were accelerated, with an apparent reduction in initial sigmoidicity. Hyperpolarizing shifts in both “a4” and isochronal inactivation “i” were also produced. KChIP2b-mediated remodeling of KV4.3 gating transitions was therefore not obligatorily dependent upon an intact N-terminus. To account for these effects we propose that KChIP2 regulatory domains exist in KV4.3 α subunit regions outside of the proximal N-terminal. In addition to regulating macroscopic inactivation, we also propose that the KV4.3 N-terminus may act as a novel regulator of deactivation-recovery coupling.
doi:10.4161/chan.5.1.13963
PMCID: PMC3052207  PMID: 21057209
KV4 channels; N-terminal domain; N-type inactivation; macroscopic inactivation and recovery; closed state inactivation and recovery; KChIP2 isoforms
6.  Variability in Pesticide Deposition and Source Contributions to Snowpack in Western US National Parks 
Environmental science & technology  2010;44(12):4452-4458.
Fifty-six seasonal snowpack samples were collected at remote alpine, sub-arctic, and arctic sites in eight Western US national parks during three consecutive years (2003–2005). Four current-use pesticides (CUPs) (dacthal (DCPA), chlorpyrifos, endosulfan, and γ-hexachlorocyclohexane (HCH)) and four historic-use pesticides (HUPs) (dieldrin, α-HCH, chlordane, and hexachlorobenzene (HCB)) were commonly measured at all sites, during all years. The mean coefficient of variation for pesticide concentrations was 15% for site replicate samples, 41% for intra-park replicate samples, and 59% for inter-annual replicate samples. The relative pesticide concentration profiles were consistent from year to year but unique for individual parks, indicating a regional source effect. HUP concentrations were well-correlated with regional cropland intensity when the effect of temperature on snow-air partitioning was considered. The mass of individual CUPs used in regions located one-day upwind of the parks was calculated using air mass back trajectories and this was used to explain the distribution of CUPs among the parks. The percent of the snowpack pesticide concentration due to regional transport was high (>75%) for the majority of pesticides in all parks. These results suggest that the majority of pesticide contamination in US national parks is due to pesticide use in North America.
doi:10.1021/es100290q
PMCID: PMC2919226  PMID: 20499934
7.  Marked Campylobacteriosis Decline after Interventions Aimed at Poultry, New Zealand 
Emerging Infectious Diseases  2011;17(6):1007-1015.
A population-level food safety response successfully reduced disease incidence.
Beginning in the 1980s, New Zealand experienced rising annual rates of campylobacteriosis that peaked in 2006. We analyzed notification, hospitalization, and other data to explore the 2007–2008 drop in campylobacteriosis incidence. Source attribution techniques based on genotyping of Campylobacter jejuni isolates from patients and environmental sources were also used to examine the decline. In 2008, the annual campylobacteriosis notification rate was 161.5/100,000 population, representing a 54% decline compared with the average annual rate of 353.8/100,000 for 2002–2006. A similar decline was seen for hospitalizations. Source attribution findings demonstrated a 74% (95% credible interval 49%–94%) reduction in the number of cases attributed to poultry. These reductions coincided with the introduction of a range of voluntary and regulatory interventions to reduce Campylobacter spp. contamination of poultry. The apparent success of these interventions may inform approaches other countries could consider to help control foodborne campylobacteriosis.
doi:10.3201/eid1706.101272
PMCID: PMC3358198  PMID: 21749761
bacteria; foodborne infections; Campylobacter; epidemiology; surveillance; poultry; food supply; bacterial typing; research; New Zealand
9.  Enhanced Psychosocial Well-Being Following Participation in a Mindfulness-Based Stress Reduction Program is Associated with Increased Natural Killer Cell Activity 
Mindfulness-based stress reduction (MBSR) programs have consistently been shown to enhance the psychosocial well-being of participants. Given the well-established association between psychosocial factors and immunologic functioning, it has been hypothesized that enhanced psychosocial well-being among MBSR participants would be associated with corresponding changes in markers of immune activity.
Objective
To examine changes in psychosocial and immunologic measures in a heterogeneous patient sample following participation in a MBSR program.
Design
A single-group, pre-test post-test design was utilized.
Setting
The intervention was conducted at an academic health center.
Subjects
This pilot study involved twenty-four participants (aged 28–72 years). Inclusion criteria were: ≥18 years of age, English-speaking, no known autoimmune disorder.
Intervention
The intervention was an 8-week MBSR program.
Outcome Measures
Distress and quality of life (QOL) measures included the Brief Symptom Inventory-18 and the Medical Outcomes Survey (MOS) Short-Form Health Survey (SF-36), respectively. Immunologic measures included natural killer (NK) cell cytolytic activity and C-reactive protein (CRP).
Results
Patients completed psychosocial assessments and provided a blood sample at baseline (pre-MBSR) and within 2 weeks post-MBSR. Significant improvements in anxiety and overall distress as well as across multiple domains of QOL were observed from baseline to post-MBSR. Reductions in anxiety and overall distress were associated with reductions in CRP. Patients who reported improvement in overall mental well-being also showed increased NK cytolytic activity from pre- to post-MBSR, whereas patients who reported no improvement in mental well-being showed no change in NK cytolytic activity.
Conclusion
Positive improvement in psychological well-being following MBSR was associated with increased NK cytolytic activity and decreased levels of CRP.
doi:10.1089/acm.2009.0018
PMCID: PMC2921566  PMID: 20455784
mindfulness-based stress reduction; psychosocial functioning; natural killer cell activity; C-reactive protein
10.  Enhanced Psychosocial Well-Being Following Participation in a Mindfulness-Based Stress Reduction Program Is Associated with Increased Natural Killer Cell Activity 
Abstract
Background
Mindfulness-based stress reduction (MBSR) programs have consistently been shown to enhance the psychosocial well-being of participants. Given the well-established association between psychosocial factors and immunologic functioning, it has been hypothesized that enhanced psychosocial well-being among MBSR participants would be associated with corresponding changes in markers of immune activity.
Objectives
The objectives of this study were to examine changes in psychosocial and immunologic measures in a heterogeneous patient sample following participation in a MBSR program.
Design
A single-group, pretest/post-test design was utilized.
Setting
The intervention was conducted at an academic health center.
Subjects
This pilot study involved 24 participants (aged 28–72 years). Inclusion criteria were as follows: ≥18 years of age, English-speaking, and no known autoimmune disorder.
Intervention
The intervention was an 8-week MBSR program.
Outcome measures
Distress and quality of life (QOL) measures included the Brief Symptom Inventory-18 and the Medical Outcomes Survey Short-Form Health Survey, respectively. Immunologic measures included natural killer (NK) cell cytolytic activity and C-reactive protein (CRP).
Results
Patients completed psychosocial assessments and provided a blood sample at baseline (pre-MBSR) and within 2 weeks post-MBSR. Significant improvements in anxiety and overall distress as well as across multiple domains of QOL were observed from baseline to post-MBSR. Reductions in anxiety and overall distress were associated with reductions in CRP. Patients who reported improvement in overall mental well-being also showed increased NK cytolytic activity from pre- to post-MBSR, whereas patients who reported no improvement in mental well-being showed no change in NK cytolytic activity.
Conclusions
Positive improvement in psychologic well-being following MBSR was associated with increased NK cytolytic activity and decreased levels of CRP.
doi:10.1089/acm.2009.0018
PMCID: PMC2921566  PMID: 20455784
11.  Chitinase-Like Proteins Are Autoantigens in a Model of Inflammation-Promoted Incipient Neoplasia 
Genes & Cancer  2011;2(1):74-87.
An important role for B cells and immunoglobulin deposition in the inflammatory tumor cell environment has been recognized in several cancers, and this is recapitulated in our murine model of inflammation-associated carcinogenesis: transgenic mice expressing the Epstein-Barr virus oncogene LMP1 in epithelia. Similarly in several autoimmune disorders, immunoglobulin deposition represents a key underlying event in the disease process. However, the autoantigens in most cases are not known. In other studies, overexpression of the enzymatically inactive mammalian chitinase-like proteins (CLPs) has been observed in a number of autoimmune disorders and numerous cancers, with expression correlated with poor prognosis, although the function of these proteins is largely unknown. We have now linked these observations demonstrating that overexpression of the CLPs renders them the targets for autoantigenicity during carcinogenic progression. We show that the CLPs, Chi3L1, Chi3L3 /YM1, and Chi3L4/YM2, are abundantly overexpressed in the transgenic epidermis at an early, preneoplastic stage and secreted into the serum. Immunoglobulin G reactive to the CLPs is detected in the serum and deposited in the hyperplastic tissue, which goes on to become inflamed and progressively displastic. The CLPs are also upregulated in chemical carcinogen-promoted lesions in both transgenic and wild-type mice. Expression of the related, active chitinases, Chit1 and AMCase, increases following infiltration of inflammatory cells. In this model, the 3 CLPs are autoantigens for the tissue-deposited immunoglobulin, which we propose plays a causative role in promoting the inflammation-associated carcinogenesis. This may reflect their normal, benign function to promote tissue remodeling and to amplify immune responses. Their induction during carcinogenesis and consequent autoantigenicity provides a missing link between the oncogenic event and subsequent inflammation. This study identifies the CLPs as important and novel therapeutic targets to limit inflammation in cancer and potentially also autoimmune disorders.
doi:10.1177/1947601911402681
PMCID: PMC3111005  PMID: 21779482
chitinase-like protein; carcinoma; inflammation; LMP1; EBV
12.  A questionnaire for determining prevalence of diabetes related foot disease (Q-DFD): construction and validation 
Background
Community based prevalence for diabetes related foot disease (DRFD) has been poorly quantified in Australian populations. The aim of this study was to develop and validate a survey tool to facilitate collection of community based prevalence data for individuals with DRFD via telephone interview.
Methods
Agreed components of DRFD were identified through an electronic literature search. Expert feedback and feedback from a population based construction sample were sought on the initial draft. Survey reliability was tested using a cohort recruited through a general practice, a hospital outpatient clinic and an outpatient podiatry clinic. Level of agreement between survey findings and either medical record or clinical assessment was evaluated.
Results
The Questionnaire for Diabetes Related Foot Disease (Q-DFD) comprised 12 questions aimed at determining presence of peripheral sensory neuropathy (PN) and peripheral vascular disease (PVD), based on self report of symptoms and/or clinical history, and self report of foot ulceration, amputation and foot deformity. Survey results for 38 from 46 participants demonstrated agreement with either clinical assessment or medical record (kappa 0.65, sensitivity 89.0%, and specificity 77.8%). Correlation for individual survey components was moderate to excellent. Inter and intrarater reliability and test re-test reliability was moderate to high for all survey domains.
Conclusion
The development of the Q-DFD provides an opportunity for ongoing collection of prevalence estimates for DRFD across Australia.
doi:10.1186/1757-1146-2-34
PMCID: PMC2789712  PMID: 19939276
13.  Novel Method for Determination of Substance P Levels in Unextracted Human Plasma by Using Acidification▿  
Substance P (SP) is a member of the tachykinin family and has an important role in immune responses. SP is detectable in plasma in a free and bound state. Simple modification of a commercially available SP enzyme-linked immunosorbent assay allows the dissociation and capture of plasma SP without solid-phase extraction.
doi:10.1128/CVI.00406-08
PMCID: PMC2668282  PMID: 19193832
14.  Perceived stress is associated with impaired T-cell response to HPV16 in women with cervical dysplasia 
Background
Infection with high-risk subtypes of human papillomavirus (HPV) is a central factor in the development of cervical neoplasia. Cell-mediated immunity against HPV16 plays an important role in the resolution of HPV infection and in controlling cervical disease progression. Research suggests that stress is associated with cervical disease progression, but few studies have examined the biological mechanisms that may be driving this association.
Purpose
This study examines whether stress is associated with immune response to HPV16 among women with cervical dysplasia.
Methods
Seventy-four women presenting for colposcopy completed measures of health behaviors, stressful life events and perceived stress (Perceived Stress Scale). A blood sample was obtained to evaluate proliferative T-cell response to HPV16, and a cervical sample was obtained during gynecologic exam for HPV-typing.
Results
Over 55% tested positive for one or more HPV subtypes. Women who did not show proliferative responses to HPV (i.e. non-responders) were more likely to be HPV+ compared to women who had a response (i.e. responders). Consistent with study hypotheses, logistic regression revealed that higher levels of perceived stress were associated with a non-response to HPV16, controlling for relevant covariates. Stressful life events were not associated with T-cell response to HPV.
Conclusions
Higher levels of perceived stress are associated with impaired HPV-specific immune response in women with cervical dysplasia, suggesting a potential mechanism by which stress may influence cervical disease progression.
doi:10.1007/s12160-007-9007-6
PMCID: PMC2396791  PMID: 18347908
stress; human papillomavirus; cervical dysplasia; T-cell proliferative response
15.  Quantifying the profile and progression of impairments, activity, participation, and quality of life in people with Parkinson disease: protocol for a prospective cohort study 
BMC Geriatrics  2009;9:2.
Background
Despite the finding that Parkinson disease (PD) occurs in more than one in every 1000 people older than 60 years, there have been few attempts to quantify how deficits in impairments, activity, participation, and quality of life progress in this debilitating condition. It is unclear which tools are most appropriate for measuring change over time in PD.
Methods and design
This protocol describes a prospective analysis of changes in impairments, activity, participation, and quality of life over a 12 month period together with an economic analysis of costs associated with PD. One-hundred participants will be included, provided they have idiopathic PD rated I-IV on the modified Hoehn & Yahr (1967) scale and fulfil the inclusion criteria. The study aims to determine which clinical and economic measures best quantify the natural history and progression of PD in a sample of people receiving services from the Victorian Comprehensive Parkinson's Program, Australia. When the data become available, the results will be expressed as baseline scores and changes over 3 months and 12 months for impairment, activity, participation, and quality of life together with a cost analysis.
Discussion
This study has the potential to identify baseline characteristics of PD for different Hoehn & Yahr stages, to determine the influence of disease duration on performance, and to calculate the costs associated with idiopathic PD. Valid clinical and economic measures for quantifying the natural history and progression of PD will also be identified.
Trial Registration
ACTRN12609000008224
doi:10.1186/1471-2318-9-2
PMCID: PMC2649133  PMID: 19152709
16.  Non-Native R1 Substitution in the S4 Domain Uniquely Alters Kv4.3 Channel Gating 
PLoS ONE  2008;3(11):e3773.
The S4 transmembrane domain in Shaker (Kv1) voltage-sensitive potassium channels has four basic residues (R1–R4) that are responsible for carrying the majority of gating charge. In Kv4 channels, however, R1 is replaced by a neutral valine at position 287. Among other differences, Kv4 channels display prominent closed state inactivation, a mechanism which is minimal in Shaker. To determine if the absence of R1 is responsible for important variation in gating characteristics between the two channel types, we introduced the V287R mutant into Kv4.3 and analyzed its effects on several voltage sensitive gating transitions. We found that the mutant increased the voltage sensitivity of steady-state activation and altered the kinetics of activation and deactivation processes. Although the kinetics of macroscopic inactivation were minimally affected, the characteristics of closed-state inactivation and recovery from open and closed inactivated states were significantly altered. The absence of R1 can only partially account for differences in the effective voltage sensitivity of gating between Shaker and Kv4.3. These results suggest that the S4 domain serves an important functional role in Kv4 channel activation and deactivation processes, and also those of closed-state inactivation and recovery.
doi:10.1371/journal.pone.0003773
PMCID: PMC2582947  PMID: 19020667
17.  Measurement of Plasma-Derived Substance P: Biological, Methodological, and Statistical Considerations▿  
Clinical and Vaccine Immunology  2006;13(11):1197-1203.
The undecapeptide substance P (SP) is a member of the tachykinin family of neurotransmitters, which has a pivotal role in the regulation of inflammatory and immune responses. One of the major barriers to the study of the in vivo role of SP in a number of immune disorders is the accurate measurement of SP in fluids. This is reflected in the variability of reported SP levels in serum and plasma of humans in both healthy and diseased states. This study was initiated in order to identify sources of variability by the comparative evaluation of the influences of sample preparation and analytical detection methods on the measurement of SP in plasma. The results indicate that sample preparation (peptide extraction versus no extraction) and the choice of analytical method for SP quantitation may yield significantly different values and may contribute to the variability in SP values reported in the literature. These results further emphasize the need for careful consideration in the selection of methods for SP quantitation, as well as caution in the interpretation and comparison of data reported in the literature.
doi:10.1128/CVI.00174-06
PMCID: PMC1656550  PMID: 16971517
22.  Measurement of Cytokine Secretion, Intracellular Protein Expression, and mRNA in Resting and Stimulated Peripheral Blood Mononuclear Cells 
Quantitation of cytokine production is a valuable adjunct to standard immunologic assays in defining several pathologic processes. Nevertheless, there is little agreement about which tissues should be assayed, which type of assay should be performed, and which stimulation protocol should be used. As these types of assays enter the clinical arena, there is need for standardization. There is also a need to maximize the amount of information which may be derived from a single sample. We compared secreted interleukin 4 (IL-4), IL-2, IL-6, tumor necrosis factor alpha (TNF-α), and gamma interferon proteins as measured by enzyme-linked immunosorbent assay with intracellular cytokine production (IL-2 and gamma interferon) as detected by flow cytometry and quantitative competitive PCR for IL-2, IL-4, TNF-α, and gamma interferon mRNA and cDNA. Results from unstimulated cells and cells stimulated with phorbol myristate acetate, phytohemagglutinin, and phorbol myristate acetate plus phytohemagglutin were compared. All three methodologies detected significant stimulation of cytokine production. The combination of phytohemagglutinin and phorbol myristate acetate was overall the most-potent stimulus.
PMCID: PMC95986  PMID: 11063499
24.  Multisite Comparison of CD4 and CD8 T-Lymphocyte Counting by Single- versus Multiple-Platform Methodologies: Evaluation of Beckman Coulter Flow-Count Fluorospheres and the tetraONE System 
New analytic methods that permit absolute CD4 and CD8 T-cell determinations to be performed entirely on the flow cytometer have the potential for improving assay precision and accuracy. In a multisite trial, we compared two different single-platform assay methods with a predicate two-color assay in which the absolute lymphocyte count was derived by conventional hematology. A two-color method employing lymphocyte light scatter gating and Beckman Coulter Flow-Count fluorospheres for absolute counting produced within-laboratory precision equivalent to that of the two-color predicate method, as measured by coefficient of variation of replicate measurements. The fully automated Beckman Coulter tetraONE System four-color assay employing CD45 lymphocyte gating, automated analysis, and absolute counting by fluorospheres resulted in a small but significant improvement in the within-laboratory precision of CD4 and CD8 cell counts and percentages suggesting that the CD45 lymphocyte gating and automated analysis might have contributed to the improved performance. Both the two-color method employing Flow-Count fluorospheres and the four-color tetraONE System provided significant and substantial improvements in between-laboratory precision of absolute counts. In some laboratories, absolute counts obtained by the single-platform methods showed small but consistent differences relative to the predicate method. Comparison of each laboratory's absolute counts with the five-laboratory median value suggested that these differences resulted from a bias in the absolute lymphocyte count obtained from the hematology instrument in some laboratories. These results demonstrate the potential for single-platform assay methods to improve within-laboratory and between-laboratory precision of CD4 and CD8 T-cell determinations compared with conventional assay methods.
PMCID: PMC95877  PMID: 10799444

Results 1-25 (42)