Unlike the collection of blood for serum, which is an invasive procedure and also requires technical expertise and disposable syringes, urine can be collected easily and frequently without causing any inconvenience to the patient

Unlike the collection of blood for serum, which is an invasive procedure and also requires technical expertise and disposable syringes, urine can be collected easily and frequently without causing any inconvenience to the patient. other than CE and 12% of urine samples from healthy controls. The circulating antigen was detected in the serum in 13 of 16 (81.25%) surgically confirmed cases, 6 of 10 (60%) ultrasound-proven cases, and 13 of 14 (92.86%) clinically diagnosed cases of CE. False-positive reactions were observed with three sera (12.5%) from controls with other parasitic diseases. The low sensitivity of Co-A for detection of antigen in the urine of a patient whose serum was positive for the antigen is possibly due to low levels of antigen in the urine. Unlike the collection of blood for serum, which is an invasive procedure and also requires technical expertise and disposable syringes, urine can be collected easily and frequently without causing any inconvenience to the patient. Urine as a clinical specimen alternative to serum would be immensely useful in the diagnosis of CE, particularly in a rural or field setting. In such situations as well as in poorly equipped laboratories, Co-A has the potential to be used as a simple, rapid, and economical slide agglutination test for detection of urinary hydatid antigen in the diagnosis of CE. Human cystic echinococcosis (CE), caused by larvae (hydatid cysts) of the dog tapeworm for 10 min at 4C. The supernatant was discarded, and the concentrated pellet of urine was resuspended in 0.1 ml of phosphate-buffered saline (PBS) (pH 7.2). Both unconcentrated and concentrated urine specimens from each patient were tested in parallel for hydatid antigen by Co-A. Hyperimmune antiserum. Hyperimmune hydatid antiserum Amotosalen hydrochloride was raised in GPIIIa rabbits as per the procedure described by us earlier (15). The antibody titer of the antiserum was 1:1,024 as measured by the indirect hemagglutination (IHA) test. The antiserum was purified as per the method described by Gottstein (4). Briefly, 1 ml of cold serum was mixed with 1 ml of cold saline at pH 7. The serum-saline mixture (2 ml) was added dropwise to 2 ml of cold saturated ammonium sulfate (pH 7) with stirring for 30 min on ice and then centrifuging at 3,000 rpm at 0C. The supernatant was discarded and the precipitate was suspended in 2 ml of saline, and the procedure was repeated until the supernatant was colorless. The final precipitate was suspended in 1 ml and dialyzed against PBS (pH 7.2) to remove all the residual ammonium sulfate. Titer of the purified antiserum was 1:2,048 by the IHA test. Co-A. The Co-A test was performed to detect hydatid antigen in the urine as per the procedure described herein. It consists of the following steps. (i) Preparation of bacterial cells. (Cowans’ strain I) bearing protein A (SAPA) was used. Amotosalen hydrochloride The cells were prepared as per the method described by Shariff and Parija (15). Briefly, cells were grown on Mueller-Hinton agar at 37C for 18 h and then were harvested, centrifuged at 3,000 for 10 min, and washed three times in PBS, pH 7.2, containing 0.05% sodium azide. The pellet was fixed in 10 volumes of 1 1.5% formaldehyde in PBS, pH 7.2, at room temperature for 90 min; washed three times in PBS, pH 7.2; Amotosalen hydrochloride resuspended to 10 volumes of buffer containing 0.05% sodium azide; and heated for 5 min at 80C. The SAPA cells were again washed twice in PBS, pH 7.2, and a 10% suspension in PBS, pH 7.2, containing 0.05% sodium azide was made. (ii) Sensitization of SAPA cells. The SAPA cells were sensitized with purified hyperimmune hydatid antiserum immediately after their preparation. One milliliter of a 10% suspension of SAPA cells was added to 0.1 ml of specific antiserum (titer, 1:2,048); these were mixed well and left at room temperature for 30 min. The cells were then washed in PBS (pH 7.2) and resuspended to a concentration of 2% in PBS (pH 7.2) containing 0.1% sodium azide. The sensitized reagent was stored at 4C. A 2% suspension of unsensitized cells was used as control. (iii) Co-A test. The test was performed on a clean slide divided with a glass-marking pen into two halves. A drop of test urine was placed on each half of the slide. An equal volume of 2% sensitized SAPA cell suspension was added to the urine on one half. The same volume of a 2% suspension of unsensitized SAPA cells was added to the urine on the other half of the slide as cell control. The slide was then rotated.