A synthetic analog of Shiga toxin (Stx) receptor (Synsorb Pk) was quantitatively assessed to determine whether it can protect human renal adenocarcinoma cells (ACHN cells) from the cytotoxicity of Stx1 and Stx2 by coincubation experiments. Coincubation of 100 and 20 ng of Stx1 and Stx2 with 50 mg of Synsorb Pk for 1 hr at 37 C in 1 ml of Eagle's Minimum Essential Medium supplemented with 1% (v/v) nonessential amino acid and 10% (v/v) fetal calf serum protected 50% of the cells from the cytotoxic effect. Chromosorb P, an inert matrix control, did not absorb the Stxs at all. Heat‐treatment (boiled for 10 min) to Synsorb Pk caused a 50% decrease in Stx2‐binding activity, but did not effect the Stx1 binding. Further, Stxs bound to Synsorb Pk could be demonstrated. When 20 mg of Synsorb Pk was coincubated for 30 min at 37 C in 1 ml of phosphate‐buffered saline with 1 and 10 ng or more of Stx1 or Stx2, respectively, the toxins could be detected on the surface when the bound toxins on Synsorb Pk were used as the solid phase in enzyme immunoassay. The amount of 100 ng/ml of both Stx1 and Stx2 appeared to saturate 20 mg/ml of Synsorb Pk after coincubating for 30 min at 37 C. While assessing the Stxs' binding activity to Synsorb Pk, it was demonstrated that Stx1 had a higher affinity to Pk trisaccharide than Stx2. These observations provide useful information on the effectiveness of Synsorb Pk to trap and eliminate free Stxs produced in the gut of patients infected by Stx‐producing Escherichia coli, and to prevent the progression of hemorrhagic colitis to hemolytic uremic syndrome.
"Quix" is an immunochromatography-based direct detection kit for the E. coli O157 LPS antigen in the patient's stool. The present study was conducted to evaluate the efficacy of the kit for rapid diagnosis of enterohemorrhagic Escherichia coli (EHEC) O157 infection. Sensitivity of the kit was determined using a pure culture of a clinical isolate of E. coli O157. Analytical sensitivity was found to be 5 x 10(5) CFU/ml. When compared with the culture method using fecal samples of 64 patients and with bloody diarrhea, sensitivity and specificity were 95.0% (19/20) and 86.4% (38/44), respectively, and overall agreement to culture method was 89.1% (57/64). One patient was found positive by culture method while negative in the present method, where the sample contained a low number of the cells less than the detection limit. Four of the six patients with a negative result by culture method and positive in the present method, were confirmed E. coli O157 infection by positive IgM antibody response against the E. coli O157 LPS. The discrepancy between the two methods seemed to be attributable to antibiotic administration. In one patient, Salmonella urbana (O30(1)30(2)) was detected. The O30(1) antigen of this bacterium is well known to be identical to the E. coli O157 antigen. When the present method was compared with an ELISA-based E. coli O157 LPS antigen detection kit, sensitivity and specificity were 100% (11/11) and 82.1% (23/28), and overall agreement to ELISA method was 87.2% (34/39). From these findings, Quix is useful as a rapid diagnostic kit in the primary clinics, outpatient or bedside use. E. coli O157 LPS antigen in patient's fecal samples can be detected in about five minutes with this simple procedure. Early diagnosis using such a simple kit will largely contribute for the early treatment and prevention of severe complications of the E. coli O157 infection.
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