Biomolecular engineering of antidehydroepiandrosterone antibodies: a new perspective in cancer diagnosis and treatment using single-chain antibody variable fragment.

AIM: To develop a monoclonal antibody against dehydroepiandrosterone (DHEA) and miniaturize it, generating a single-chain antibody variable fragment (scFv) against DHEA as an adrenocortical carcinoma (ACC) marker. MATERIAL & METHODS: DHEA conjugated to keyhole limpet hemocyanin was used as an immunogen to obtain anti-DHEA hybridomas. Variable fragments were cloned from hybridoma 5B7 total RNA, and used to detect DHEA in normal adrenal tissue and ACC cells. RESULTS: IgM monoclonal antibody was highly specific, and the recombinant scFv preserved parental antibody characteristics, allowing tissue localization of DHEA. CONCLUSION: Undefined small lesions are challenges for clinicians and impact clinical adrenocortical tumor management. Generating an anti-DHEA scFv facilitates development of imaging tests for early diagnosis of pediatric ACC.

Immunodetection of the "brown" spider (Loxosceles intermedia) dermonecrotoxin with an scFv-alkaline phosphatase fusion protein.

Bites by spiders from Loxosceles genus often lead to a wide variance in envenomation profile of patients and diagnosis is difficult due to the number of diseases that mimic loxoscelism. In such a context, it is of interest to consider the design of standardized recombinant colorimetric antibodies for diagnosis and specific detection of individual circulating toxins in biological fluids of envenomed patients. We have previously prepared a monoclonal murine IgG (LiMab7) that reacts with Loxosceles intermedia venom components of 32-35kDa and neutralizes the dermonecrotic activity of the venom. Here, we re-engineered LiMab7 into a colorimetric bifunctional protein consisting in the corresponding single-chain antibody fragment (scFv) fused to alkaline phosphatase (AP) of Escherichia coli. The immune tracer was tested in two different types of immunoassays and it proved to be efficient in both. Thus, this recombinant fusion protein (scFv-LiMab7/AP) can be used for rapid and specific immunotitration of L. intermedia venom with a linear range of 39-20000ng/mL and a detection limit of 39ng/mL without any cross-reaction.

Left atrial and left ventricular diastolic function in chronic Chagas disease.

BACKGROUND: Left atrial (LA) and left ventricular (LV) diastolic function analysis can yield new strategies to recognize early cardiac involvement and prognostic indicators in Chagas disease. METHODS: Patients with Chagas disease with the indeterminate (n = 69) or with the cardiac form (32 with changes limited to electrocardiography [stage A], 25 with changes in LV systolic function but no heart failure [HF; stage B], and 26 with HF) underwent evaluation of LV diastolic function (mitral inflow, pulmonary vein flow, color M-mode echocardiography, and tissue Doppler analysis), and LA function by three-dimensional echocardiography and strain analysis and were prospectively followed for the occurrence of clinical events. Echocardiograms were also obtained from 32 controls. RESULTS: LV diastolic dysfunction was gradually more prevalent and severe across groups from patients with the indeterminate form of Chagas disease to patients with HF. Tissue Doppler was the best tool to demonstrate the worsening of LV diastolic function across the groups (E' velocity: controls, 12.6 ± 2.3 cm/sec; patients with the indeterminate form, 12.1 ± 3.1 cm/sec; stage A, 10.3 ± 2.9 cm/sec; stage B, 8.3 ± 2.8 cm/sec; patients with HF, 5.6 ± 1.9; P < .0001). Although maximum LA volume was increased only in patients with HF, minimum LA volume (controls, 8 ± 2 mL/m(2); patients with the indeterminate form, 8 ± 2 mL/m(2); stage A, 9 ± 3 mL/m(2); stage B, 11 ± 4 mL/m(2); patients with HF, 27 ± 17 mL/m(2); P < .0001) and precontraction LA volume (controls, 11 ± 3 mL/m(2); patients with the indeterminate form, 12 ± 3 mL/m(2); stage A, 13 ± 4 mL/m(2); stage B, 16 ± 5 mL/m(2); patients with HF, 32 ± 19 mL/m(2); P < .0001) were increased in all cardiac form groups. LA conductive function was depressed in all cardiac form groups, while LA contractile function was depressed only in patients with HF. Cox proportional-hazards regression analysis revealed that end-systolic LV diameter (hazard ratio, 1.6; 95% confidence interval, 0.9-2.8; P = .09), E' velocity (hazard ratio, 0.5; 95% confidence interval, 0.3-0.8; P = .001), and peak negative global LA strain (hazard ratio, 1.21; 95% confidence interval, 1.02-1.4; P = .03), were independent predictors of clinical events. CONCLUSIONS: LV diastolic dysfunction was found in all forms of chronic Chagas disease, including those without LV systolic dysfunction. LV diastolic dysfunction may contribute to changes in LA volume and conductive function found in early stages of the cardiac form. Both LV diastolic function and LA contractile function were independent predictors of clinical events.