Microfluidic integration for automated targeted proteomic assays.

A dearth of protein isoform-based clinical diagnostics currently hinders advances in personalized medicine. A well-organized protein biomarker validation process that includes facile measurement of protein isoforms would accelerate development of effective protein-based diagnostics. Toward scalable protein isoform analysis, we introduce a microfluidic "single-channel, multistage" immunoblotting strategy. The multistep assay performs all immunoblotting steps: separation, immobilization of resolved proteins, antibody probing of immobilized proteins, and all interim wash steps. Programmable, low-dispersion electrophoretic transport obviates the need for pumps and valves. A three-dimensional bulk photoreactive hydrogel eliminates manual blotting. In addition to simplified operation and interfacing, directed electrophoretic transport through our 3D nanoporous reactive hydrogel yields superior performance over the state-of-the-art in enhanced capture efficiency (on par with membrane electroblotting) and sparing consumption of reagents (ca. 1 ng antibody), as supported by empirical and by scaling analyses. We apply our fully integrated microfluidic assay to protein measurements of endogenous prostate specific antigen isoforms in (i) minimally processed human prostate cancer cell lysate (1.1 pg limit of detection) and (ii) crude sera from metastatic prostate cancer patients. The single-instrument functionality establishes a scalable microfluidic framework for high-throughput targeted proteomics, as is relevant to personalized medicine through robust protein biomarker verification, systematic characterization of new antibody probes for functional proteomics, and, more broadly, to characterization of human biospecimen repositories.

Use of orlistat (xenical) to treat chylous ascites.

Chylous ascites is uncommon and occurs in about 1 in 20,000 hospital admissions. Causes include disruption of the lymphatic system due to malignancy, cirrhosis, surgery, or radiation therapy. The mainstay of therapy has been low-fat diet supplemented with medium-chain triglyceride oil. However, dietary compliance can be difficult to achieve for adequate response. We report a 47-year-old man with hepatitis C and alcohol-related cirrhosis with new-onset chylous ascites and chylothorax. His ascites triglyceride was 585 mg/dL, and the pleural fluid triglyceride was 691 mg/dL. Ascitic and pleural fluid cytology and acid-fast bacilli stain were negative. The patient was treated with low-fat diet and medium-chain triglyceride oil. However, his ascites remained chylous after 1 week of treatment because of poor compliance with the dietary restrictions. Orlistat was then added to his treatment regimen. A half week later, the chylous component of his ascites resolved. Remaining high-volume clear ascites was treated with placement of a transjugular intrahepatic portosystemic shunt. To our knowledge, orlistat has never been used in the treatment of chylous ascites. This case suggests the potential value of adding orlistat to low-fat diet and medium-chain triglyceride oil in the treatment of chylous ascites, especially in patients who are unable to comply with the dietary restrictions.

Cytomegalovirus ischemic colitis: a near-fatal presentation of HIV infection.

Cytomegalovirus infection occurs in immunocompromised patients. We present a 45-year-old male with no prior medical history who presented to the hospital with weight loss and non-bloody diarrhea. During hospitalization, he developed severe hematochezia and hypotension. Colonoscopy revealed dusky, friable mucosa. The patient arrested and was resuscitated. Specimen from emergent colectomy showed ischemic changes secondary to cytomegalovirus infection of endothelium and small-vessel thrombosis. An HIV test was subsequently positive with CD4 count of 2 per microliter. The patient was treated with antiretroviral therapy and ganciclovir. He survived postoperative infections and was eventually discharged. In summary, this case of near-fatal cytomegalovirus colitis represents an unusual presentation of undiagnosed HIV infection. Cytomegalovirus infection should be included in the differential diagnosis of immunocompromised patients with gastrointestinal symptoms. Hematochezia may be from intestinal ulceration or severe ischemic damage. Antiretroviral therapy and ganciclovir or foscarnet should be initiated promptly. Surgery is indicated in life-threatening hemorrhage or obvious bowel necrosis.

Is it cost-effective to treat recurrent hepatitis C infection in orthotopic liver transplantation patients?

Hepatitis C virus (HCV) recurs in the allograft almost universally after orthotopic liver transplantation (OLT), with a variable course ranging from mild hepatitis to frank cirrhosis. The uncertain prognosis after OLT has lead to widely increased use of antiviral therapy in the post-OLT setting. We compared two scenarios (antiviral therapy versus no antiviral therapy) using a Markov-based decision analytic model to simulate costs and health outcomes for recurrent HCV in three age and sex cohorts of post-OLT patients. Efficacy outcomes included total costs, cases of cirrhosis prevented, cases of death prevented, life-years gained, and cost per life-year saved. One-way sensitivity analyses were performed for sustained viral response; annual drug cost, discount rate, compliance, cirrhosis rate, decompensation rate, and cost of dying. Two-way sensitivity analyses were performed to compare effects of (1) changing sustained viral response and antiviral therapy costs, and (2) changing the sustained viral response and cirrhosis rate. The incremental cost-effectiveness ratio for the reference patient cohort of 1,000 men aged 55 years was $29,100 per life-year saved. The model was sensitive to drug costs, cirrhosis rate, and sustained viral response. The two-way sensitivity analysis showed that antiviral therapy remained cost-effective even if drug costs increased, as long as these increases were associated with higher sustained viral responses. The cost-effectiveness ratio also was sensitive to age and sex of cohort. The decision to treat HCV after OLT with antiviral therapy usually is based on many considerations. Such treatment can be cost-effective if baseline assumptions are met. Our model was sensitive to antiviral drug costs, cirrhosis rate, and sustained viral response. Patients with a progressive course of recurrent HCV are likely to have the greatest gain from antiviral therapy.