Elevated receptor for activated C kinase 1 expression is involved in intracellular Ca2+ influx and potentially associated with compromised regulatory T cell function in patients with asthma.

BACKGROUND: Regulatory T cells (T(regs)) are activated during anergy in response to T cell receptor (TCR) activation and functional immune suppression. Anergy of paediatric T(regs) is partially dependent on intracellular calcium mobility; following TCR activation, T(regs) do not exhibit increased intracellular Ca(2+) concentration ([Ca(2+) ](i)). OBJECTIVE: We determined whether [Ca(2+) ](i) in adult T(regs) defined their anergy, if intracellular Ca(2+) movement was linked to regulatory functions, whether [Ca(2+)](i) was indicative of asthma pathology, and the potential molecular mechanism responsible for Ca(2+) movement in T(regs). METHODS: T(regs) were purified by the magnetic bead method, and their regulatory functions were assessed by monitoring carboxyfluorescein succinimidyl ester-labelled responder T cell proliferation. The Ca(2+) response of Fura-2-labelled cells was measured using a video image analysis system. To analyse the functions of T(regs) at the molecular level, we generated Jurkat Tet-On(®) clones with doxycycline (Dox)-induced forkhead box P3 (FOXP3) protein expression. RESULTS: CD4(+) CD25(+) CD127(-/low) T(regs) from participants without asthma did not elicit Ca(2+) influx in response to TCR activation, exhibited little proliferation and suppressed proliferation of CD4(+) CD25(-) T cells. In contrast, under similar conditions, T(regs) from patients with asthma exhibited increased [Ca(2+)](i) and robust proliferation with partial loss of regulatory functions. FOXP3 protein levels in Tet-On(®) clones were high after both 2- and 5-day Dox treatment; however, 5-day cells were comparable with T(regs) from patients with asthma, whereas 2-day cells were similar to T(regs) from participants without asthma. Increasing [Ca(2+)](i) induced a high level of receptor for activated C kinase 1 (RACK1) expression in 5-day cells. CONCLUSIONS AND CLINICAL RELEVANCE: We confirmed that T(regs) in patients with asthma are functionally impaired and that the abnormal regulatory functions of these cells can be analysed by [Ca(2+)](i) following TCR engagement. Furthermore, the impaired functioning of T(regs) evident in patients with asthma may be due to a high level of RACK1.

Close relationship between T helper (Th)17 and Th2 response in murine allergic contact dermatitis.

BACKGROUND: Repeated exposure to allergens induces chronic allergic lesions in the skin and a shift in the cutaneous cytokine milieu to T helper (Th)2. AIM: To assess the relationships between Th17 and Th2 response during allergic contact dermatitis (ACD) in mice. METHODS: ACD was induced in C57BL/6 mice by single or repeated epicutaneous challenge of 2,4,6-trinitro-1-chlorobenzene. Relationships between Th17 and Th2 response were analyzed by immunohistochemical observations and activity of cytokines on days 8 (first challenge), 18 (11th challenge), 28 (21st challenge) and 38 (31st challenge). RESULTS: On day 8, tissue levels of interleukin (IL)-17 and IL-22 were high, whereas tissue levels of IL-4 and serum IgE concentration were low. Following acute contact dermatitis, mice developed chronic eczematous lesions on day 18, and gradually improved on days 28 and 38. Tissue IL-4 and serum IgE levels corresponded to the development and improvement of chronic eczematous lesions. Numbers of Th17 cells and tissue levels of IL-17 and IL-22 rapidly decreased as IL-4 and IgE levels increased on day 18. As levels of IL-4 and IgE decreased, the number of Th17 cells and tissue levels of IL-17 and IL-22 increased again on days 28 and 38. On day 18, tissue levels of Th17 response-inducing cytokines (IL-6, IL-23 and transforming growth factor-ß) were high, and IL-23-expressing cells appeared in abundance, when Th2 response was extremely high. IL-17 injection decreased tissue IL-4 and serum IgE levels. CONCLUSIONS: Th17 correlates closely with Th2 in murine chronic ACD induced by repeated epicutaneous challenge.

Optimal antidiarrhea treatment for antitumor agent irinotecan hydrochloride (CPT-11)-induced delayed diarrhea.

PURPOSE: An antitumor camptothecin derivative CPT-11 has proven a broad spectrum of solid tumor malignancy, but its severe diarrhea has often limited its more widespread use. We have demonstrated from a rat model that intestinal beta-glucuronidase may play a key role in the development of CPT-11-induced delayed diarrhea by the deconjugation of the luminal SN-38 glucuronide, and the elimination of the intestinal microflora by antibiotics or dosing of TJ-14, a Kampo medicine that contains beta-glucuronidase inhibitor baicalin, exerted a protective effect. In the present study, we assessed the efficacy of several potential treatments in our rat model to clarify which is the most promising treatment for CPT-11-induced delayed diarrhea. METHODS AND RESULTS: Oral dosing (twice daily from days -1 to 4) of streptomycin 20 mg/kg and penicillin 10 mg/kg (Str/Pen), neomycin 20 mg/kg and bacitracin 10 mg/kg (Neo/Bac), both of which inhibited almost completely the fecal beta-glucuronidase activity, or TJ-14 1,000 mg/kg improved the decrease in body weight and the delayed diarrhea symptoms induced by CPT-11 (60 mg/kg i.v. from days 1 to 4) to a similar extent. The efficacy was less but significant in activated charcoal (1,000 mg/kg p.o. twice daily from days -1 to 4). In a separate experiment using rats bearing breast cancer (Walker 256-TC), TJ-14, Neo/Bac, and charcoal at the same dose regimen improved CPT-11-induced intestinal toxicity without reducing CPT-11's antitumor activity. In contrast, oral dosing (twice a day) of cyclosporin A (50 mg/kg), a P-glycoprotein and cMOAT/MRP2 inhibitor or valproic acid (200 mg/kg), a UDP-glucuronosyltranferase inhibitor, exacerbated the intestinal toxicity without modifying CPT-11's antitumor activity. CONCLUSIONS: The result clearly demonstrated the ability of Neo/Bac, Str/Pen, and TJ-14, less but significant ability of activated charcoal, to ameliorate CPT-11-induced delayed-onset diarrhea, suggesting the treatments decreasing the exposure of the intestines to the luminal SN-38 are valuable for improvement of CPT-11-induced intestinal toxicity. In contrast, the treatments affecting the biliary excretion of CPT-11 and its metabolites might have undesirable results.

Involvement of beta-glucuronidase in intestinal microflora in the intestinal toxicity of the antitumor camptothecin derivative irinotecan hydrochloride (CPT-11) in rats.

Irinotecan hydrochloride (CPT-11), an antitumor camptothecin derivative, causes severe forms of diarrhea clinically. We characterized CPT-11-induced diarrhea histologically and enzymologically and assessed the relationships between intestinal toxicity and the activity of the enzymes that play a key role in the major metabolic pathway of CPT-11 in rats. CPT-11 (60 mg/kg i.v. for 4 days) induced intestinal toxicity characterized by severe chronic diarrhea, loss of body weight, and anorexia. Histological damage was most severe in the cecum. The segmental difference in the degree of the damage showed good correlation with the beta-glucuronidase activity in the contents of the lumen in each case, but not with the intestinal tissue carboxylesterase activity, which converts CPT-11 to its active form (7-ethyl-10-hydroxycamptothecin). Inhibition of the beta-glucuronidase activity in the intestinal microflora by antibiotics (1 mg penicillin and 2 mg streptomycin per ml of drinking water) markedly ameliorated the diarrhea and reduced cecal damage. Analysis of CPT-11 and its metabolites in the feces indicated that antibiotics completely inhibited the deconjugation of the glucuronic conjugate of 7-ethyl-10-hydroxycamptothecin by beta-glucuronidase. It is suggested that CPT-11-induced diarrhea would be attributable to the damage to the cecum, and that the inhibition of the beta-glucuronidase activity in the intestinal microflora is a major protective effect of antibiotics.

Molecular cloning of two glutamate transporter subtypes from mouse brain.

The physiological action of glutamate is terminated by diffusional processes with its subsequent removal by high-affinity transport systems localized to glial and/or neuronal elements. Several cDNAs encoding glutamate transporters have been isolated from mammalian tissues. Here, we screened a cDNA library derived from mouse cerebellum and isolated the two glutamate transporters, termed MGLT1 and MEAAC1. The MGLT1 and MEAAC1 cDNAs encode proteins of 572 and 523 amino acids, respectively. MGLT1 has 93.9% amino acid sequence identity with the rat GLT1 and MEAAC1 has 89.3% identity with the rabbit EAAC1. MEAAC1 mRNA was expressed in brain, lung, kidney and skeletal muscle, whereas expression of MGLT1 was restricted to brain.

Crystallization and characterization of a lectin obtained from a mushroom, Aleuria aurantia.

A fucose-specific lectin with a unique sugar recognizing property was purified from an orange peel mushroom, Aleuria aurantia, by using a specific affinity adsorbent prepared from L-fucose and starch. From 100 g of fruiting bodies, 145 mg of pure lectin was obtained. The lectin was crystallized and the crystals showed hexagonal bipyramid in shape. Distribution of hydrophobic and hydrophilic regions in the molecule of this lectin was predicted from the amino acid sequence deduced from the previously reported nucleotide sequence of the lectin cDNA. Circular dichroism spectra revealed a very low content of alpha-helical and beta-sheet structures and a relatively high content of turns in this lectin. From the spectrum observed in the presence of L-fucose, a hapten sugar of this lectin, certain conformational change was assumed to occur.

Development of endometrial cancer following radiation therapy for cervical carcinoma.

The clinical and histologic findings in five cases of endometrial cancer, which developed following radiation therapy for squamous cell carcinoma of the cervix, are described. The mean age at endometrial cancer diagnosis was 69 years and average latency period from initial therapy to development of endometrial carcinoma was 13.4 years. For endometrial cancer, one patient had Stage Ib, one patient had Stage IIIa, two patients had Stage IIIc, and one patient did not undergo laparotomy. The histological types were carcinosarcoma in two patients, endometrioid adenocarcinoma, grade 3 in one patient, and clear cell carcinoma in one patient. All patients died of disease within 33 months of diagnosis. Endometrial cancers that develop after radiation treatment have a preponderance of high-risk histological subtypes, and consequently have a poor prognosis. Long-term follow-up should be mandatory for patients surviving radiation therapy for cervical cancer in order to detect and effectively treat second malignancies.

Proliferative properties of human umbilical cord blood megakaryocyte progenitor cells to human thrombopoietin.

We evaluated the effects of human thrombopoietin (TPO) alone, or in combination with other several hematopoietic factors, on megakaryocyte colony growth from human umbilical cord blood CD34+ cells in serum-depleted agar cultures. The addition of TPO alone had a concentration-dependent effect on the number of pure megakaryocyte colonies and of megakaryocytes per colony (colony size). The maximally stimulating concentration of TPO generated a greater number of megakaryocyte colonies and larger megakaryocyte colony size compared with the stimulation observed with an optimal concentration of human interleukin-3 (IL-3) or stem cell factor (SCF). At the high concentration of TPO that yielded the maximum colony numbers, a substantial proportion of megakaryocyte colonies contained 100 or more cells per colony. The combination of TPO plus SCF resulted in a synergistic enhancement of both the number and size of megakaryocyte colonies. Among the combinations of TPO plus other cytokines tested, IL-3 plus TPO had a modest effect on megakaryocyte colony numbers. The generation of megakaryocyte colonies from subpopulations of CD34+ cells was further examined. The addition of TPO alone induced a greater number of megakaryocyte colonies from CD34+CD41+ cells compared with CD34+CD41- cells, and TPO plus IL-3 exerted a synergistic effect on the number of megakaryocyte colonies only from CD34+CD41- cells. In contrast to the effects on colony numbers, TPO induced larger megakaryocyte colony sizes from CD34+CD41- cells, compared with CD34+CD41+ cells. In the case of HLA-DR expression, TPO and IL-3, administered singly or in combination, induced similar megakaryocyte colony numbers and sizes from CD34+DR+ and CD34+DR- subpopulations. Ploidy analysis revealed that the majority of megakaryocytes generated from cord blood CD34+ cells in serum-free liquid cultures containing TPO displayed 2N and 4N values, suggesting that they were immature. These results indicate that, compared with IL-3 and SCF, TPO has more potent proliferative effect on human cord blood megakaryocyte progenitors, leading to greater numbers of megakaryocyte progenitors triggered for both growth and cell division, and synergizes with SCF to enhance megakaryocyte colony growth.

Action of thrombopoietin at the megakaryocyte progenitor level is critical for the subsequent proplatelet production.

Formation of proplatelets from megakaryocytes is believed to be the first step of platelet production in vitro. In this study, we evaluated the effects of recombinant human thrombopoietin (hTPO) on the development of proplatelets from a GpIIb/IIIa+ population of rat bone marrow cells highly enriched for late megakaryocyte progenitors (GpIIb/IIIa+ CFU-MK) that we recently found to be a primary target population of TPO. Quantitative measurement of hTPO-induced proplatelet formation was performed in liquid cultures. Proplatelet formation from megakaryocytes derived from GpIIb/IIIa+ CFU-MK in the presence of hTPO began on day 4 of culture and peaked the following day. On day 5 of culture, lower concentrations of hTPO expanded the number of megakaryocytes, increased the number of proplatelets and the percentage of proplatelet-developing megakaryocytes. Increasing hTPO concentrations resulted in a modest decrease in proplatelet development. We next used hTPO to derive immature or mature megakaryocytes from GpIIb/IIIa+ CFU-MK. These populations of cultured megakaryocytes spontaneously formed proplatelets when recultured in the absence of exogenous hTPO. The addition of hTPO at higher concentrations modestly augmented proplatelet production from immature megakaryocytes derived from 2-day liquid cultures. However, either murine interleukin-6 (IL-6) or human IL-11, but not rat IL-3, was more potent than hTPO in augmenting proplatelet formation from immature megakaryocytes. Each of these four cytokines had an inhibitory effect on proplatelet formation from more differentiated megakaryocytes derived from 3-day liquid cultures. These results indicate that TPO enhances proplatelet production primarily by stimulating CFU-MK to increase the number of proplatelet-forming megakaryocytes and that its action is clearly different from those of other cytokines that also stimulate megakaryocytopoiesis.

GpIIb/IIIa+ subpopulation of rat megakaryocyte progenitor cells exhibits high responsiveness to human thrombopoietin.

The recently cloned factor thrombopoietin (TPO) has been shown to exhibit megakaryocyte colony-stimulating activity in vitro. In this investigation, to further evaluate the action of TPO on megakaryocyte progenitor cells (colony-forming units-megakaryocyte [CFU-MK]), GpIIb/IIIa+ and GpIIb/IIIa- populations of CFU-MK were prepared from rat bone marrow cells based on their reactivity with P55 antibody, a monoclonal antibody against rat GpIIb/IIIa, and their responsiveness to recombinant human TPO (rhTPO) and recombinant rat interleukin-3 (rrIL-3) was examined using a megakaryocyte colony-forming assay (Meg-CSA). rhTPO supported only megakaryocyte colony growth from both fractions in a dose-dependent fashion. The mean colony size observed with the GpIIb/IIIa+ population was smaller than that seen with the GpIIb/IIIa- population. With the optimal concentration of either rhTPO or rrIL-3, similar numbers of megakaryocyte colonies were formed from the GpIIb/IIIa+ population previously shown to be highly enriched for CFU-MK. In contrast, the maximum number of megakaryocyte colonies from the GpIIb/IIIa- population stimulated by rhTPO was only 24.2% of that achieved with rrIL-3. Morphologic analysis of rhTPO-promoted megakaryocyte colonies from the GpIIb/IIIa+ population showed that the average colony size was smaller but that the mean diameter of individual megakaryocytes was larger than in megakaryocyte colonies promoted with rrIL-3. rhTPO plus rrIL-3, each at suboptimal concentrations, had an additive effect on proliferation of CFU-MK in the GpIIb/IIIa+ fraction, whereas rhTPO plus murine IL-6 or murine granulocyte-macrophage colony-stimulating factor (mG-M-CSF) modestly but significantly reduced megakaryocyte colony growth. These results indicate that TPO preferentially acts on GpIIb/IIIa+ late CFU-MK with lower proliferative capacity and interacts with some other cytokines in CFU-MK development.

Expression of adhesion molecules on cytoplasmic processes of human megakaryocytes.

Megakaryocytes generate cytoplasmic processes (CP) that penetrate endothelial cells in the bone marrow sinus, and these processes may release platelets into the circulation at their terminal stage. Adhesion between the CP and endothelial cells may be important during the extension of CP. We examined the expression of adhesion molecules of the integrin family (CDw49b, CDw49d, CDw49e, CDw49f, CD18, CD11a CD11c, and CD11b), the immunoglobulin superfamily (CD54, CD56, CD58, and CD31), the selectin family (ELAM-1, LECAM-1, and CD62), and CD44, CD41b, and CD42b on platelets, megakaryocytes, and megakaryocytes with CP. No specific adhesion molecules were observed on the megakaryocytes with CP. Three staining patterns of adhesion molecules-homogeneous, speckled, and accumulated-were observed on the megakaryocytes with CP, but not on those without CP. Platelet integrins (i.e., CD41a, CDw49b, CDw49e and CDw49f) and GPIb (CD42b) were strongly and homogeneously stained on the CP. GMP-140 CD62) was weakly stained, in a speckled pattern. CD31 (PECAM-1) was also weakly stained but accumulated selectively on the tip of the CP. ANTI-CD31 suppressed CP formation of megakaryocytes. We speculate that the homodimerization of CD31 expressed on the tips of CP and endothelial cells is important for the extension of the processes and for the migration of megakaryocytes.

Cloning and expression of a functional fucose-specific lectin from an orange peel mushroom, Aleuria aurantia.

Aleuria aurantia lectin (AAL) shows sugar-binding specificity for L-fucose. A lambda gt11 expression library was constructed from A. aurantia poly(A) RNA and screened with a polyclonal antiserum directed against AAL. An immunopositive clone carrying 1.3-kb EcoRI fragment was obtained. The fragment encoded AAL, but lacked a nucleotide sequence corresponding to the two amino-terminal amino acids. The 5'-terminal part of the fragment was replaced with a chemically synthesized DNA fragment and inserted into an expression vector to yield a plasmid pKA-1. Escherichia coli carrying pKA-1 expressed functional AAL and the recombinant AAL showed the same immunological properties as those of natural AAL.

Quantitative analysis of thrombopoietin receptors on human megakaryocytes.

Thrombopoietin (TPO), or c-MPL ligand, is the primary regulator of megakaryocyte and platelet production. TPO receptors expressed on human megakaryocytes derived from peripheral blood (PB) and cord blood (CB) progenitors cultured in the presence of TPO have now been analyzed quantitatively. Like those on human PB platelets, TPO receptors on the cultured megakaryocytes exhibited a molecular mass of approximately 80 kDa. Various characteristics of PB- and CB-derived megakaryocytes indicated that the former were more mature than the latter. Both PB- and CB-derived megakaryocytes expressed a single class of high-affinity TPO receptors, with 1933 +/- 772 (n = 3) and 184 +/- 48 (n = 4) sites per cell, respectively. These data indicate that the number of TPO receptors on human megakaryocytes increases with cell maturation.

Differential sensitivity to hydroxyl radicals of pre- and postjunctional neurovascular transmission in the isolated canine mesenteric vein.

In some pathophysiological conditions, the first target of reactive oxygen intermediates is the vascular system. Superoxide anions, when generated in the vascular circulation, may then escape into the extracellular space via an anion channel and, following dismutation to hydrogen peroxide (H(2)O(2)), form hydroxyl radicals (HO(*)). In an attempt to understand the role of HO(*) in the regulation of transmission at the sympathetic neurovascular junction, the effect of HO(*) at nerve terminals was examined by measuring the amount of noradrenaline (NA) released from isolated, spirally cut, superfused canine mesenteric vein during basal and electrical stimulation (ES; 5Hz, 2ms, 9V); tension development evoked by ES was also recorded simultaneously. HO(*) was generated from Fenton's reagent (1. 5x10(-4)M H(2)O(2) plus 10(-4)M FeSO(4)); generation of HO(*) from H(2)O(2)/FeSO(4) in the superfusate was monitored by electron spin resonance spectroscopy using the spin-trap 5, 5-dimethyl-1-pyrroline-N-oxide throughout the experimental time course. Exposure to HO(*) of the helical strips produced an irreversible decrease in tension development evoked by ES with no effect on NA release, suggesting that the observed effect is elicited postjunctionally. The susceptibility of the processes of NA-mediated contraction to HO(*) may differ greatly from that of the NA release mechanism at the prejunctional site. Exposure of the strip preparation to HO(*) leads to a substantial stimulation of basal release of NA without affecting ES-evoked NA release, possibly due to enhanced non-exocytotic Ca(2+)-independent release elicited by HO(*). A direct demonstration of this concept was obtained by showing a significant increase in the basal response of NA release in Ca(2+)-free solution. The major conclusion of the present study is that HO(*) can damage NA-mediated contraction of the vascular preparations at the postjunctional site, and may selectively induce a non-exocytotic release of NA from the prejunctional site of sympathetic neurotransmission.

A novel mutation Gly 1672-->Arg in type 2A and a homozygous mutation in type 2B von Willebrand disease.

Genetic materials from 16 unrelated Japanese patients with von Willebrand disease (vWD) were analyzed for mutations. Exon 28 of the von Willebrand factor (vWF) gene, where point mutations have been found most frequent, was screened by various restriction-enzyme analyses. Six patients were observed to have abnormal restriction patterns. By sequence analyses of the polymerase chain-reaction products, we identified a homozygous R1308C missense mutation in a patient with type 2B vWD; R1597W, R1597Q, G1609R and G1672R missense mutations in five patients with type 2A; and a G1659ter nonsense mutation in a patient with type 3 vWD. The G1672R was a novel missense mutation of the carboxyl-terminal end of the A2 domain. In addition, we detected an A/C polymorphism at nucleotide 4915 with HaeIII. There was no particular linkage disequilibrium of the A/C polymorphism, either with the G/A polymorphism at nucleotide 4391 detected with Hphl or with the C/T at 4891 detected with BstEII.

Primary structure of a fucose-specific lectin obtained from a mushroom, Aleuria aurantia.

Aleuria aurantia lectin (AAL) is a protein composed of two identical subunits having no carbohydrate chain and shows sugar-binding specificity for L-fucose. Full-length cDNA encoding for the lectin has been isolated from a lambda gt11 library, screened with an antiserum directed against AAL. The cDNA clone contained 1,370 nucleotides and an open reading frame of 939 nucleotides encoding 313 amino acids. The amino-terminal sequence (residues 1-30) of the lectin isolated from the mushroom coincided with the deduced amino acid sequence starting from proline at the 2nd residue, indicating that the mature AAL consists of 312 amino acids. Its molecular weight is calculated to be 33,398. The deduced amino acid sequence shows that AAL includes six internal homologous regions, and has considerable homology with a hemagglutinin from a Gram-negative bacterium, Myxococcus xanthus, which forms a fruiting body. No significant homology was observed with higher plant or animal lectins. The recombinant AAL produced by Escherichia coli JM109 carrying the AAL expression plasmid pKA-1 [Fukumori, F. et al. (1989) FEBS Lett. 250, 153-156] was purified from the cell lysate by affinity chromatography using a fucose-starch column, and hundreds of milligrams of the lectin was obtained. The recombinant lectin showed the same biochemical characteristics and sugar binding specificity as did the natural AAL.

Evaluation of a new amplified enzyme immunoassay (EIA) for the detection of Chlamydia trachomatis in male urine, female endocervical swab, and patient obtained vaginal swab specimens.

AIMS: To compare the performance of a new generation dual amplified enzyme immunoassay (EIA) with a molecular method for the diagnosis of Chlamydia trachomatis, using a range of urogenital samples, and to assess the reliability of testing self collected vaginal specimens compared with clinician collected vaginal specimens. METHODS: Two population groups were tested. For the first population group, first void urine samples were collected from 193 male patients with urethritis, and endocervical swabs were collected from 187 high risk commercial sex workers. All urine and endocervical specimens were tested by a conventional assay (IDEIA chlamydia), a new generation amplified immunoassay (IDEIA PCE chlamydia), and the Amplicor polymerase chain reaction (PCR). Discrepant results obtained among the three sample types were confirmed using a nested PCR test with a different plasmid target region. For the second population group, four swab specimens, including one patient obtained vaginal swab, two clinician obtained endocervical swabs, and one clinician obtained vaginal swab, were collected from 91 high risk sex workers. Self collected and clinician collected vaginal swabs were tested by IDEIA PCE chlamydia. Clinician obtained endocervical swabs were assayed by IDEIA PCE chlamydia and Amplicor PCR. RESULTS: The performance of the IDEIA PCE chlamydia test was comparable to that of the Amplicor PCR test when male urine and female endocervical swab specimens were analysed. The relative sensitivities of IDEIA, IDEIA PCE, and Amplicor PCR on male first void urine specimens were 79.3%, 91.4%, and 100%, respectively. The relative sensitivities of the three tests on female endocervical specimens were 85.0%, 95.0%, and 100%, respectively. The positivity rates for patient collected vaginal specimens and clinician collected vaginal specimens by IDEIA PCE were 25.2% and 23.1%, respectively, whereas those for clinician collected endocervical swabs by PCR and IDEIA PCE were both 27.5%. CONCLUSIONS: IDEIA PCE chlamydia is a lower cost but sensitive alternative test to PCR for testing male urine samples and female endocervical swabs. In addition, self collected or clinician collected vaginal specimens tested by IDEIA PCE chlamydia are a reliable alternative to analysing endocervical specimens.