All sections counterstained with hematoxylin

  • by

All sections counterstained with hematoxylin. median 22.5 months) and two did not improve after the procedure. Conclusions: In young men with Ma2-encephalitis, 1) the disorder should be attributed to a germ-cell neoplasm of the testis unless another Ma2-expressing tumor is found, 2) negative tumor markers, ultrasound, body CT, or PET do not exclude an intratubular germ-cell neoplasm of the testis, and 3) if no tumor is found, the presence of the five indicated criteria should prompt consideration of orchiectomy. Tumors involved in paraneoplastic neurologic disorders are usually small, confined to a specific organ, or detectable at the organ-draining lymph nodes.1,2 Recent studies show that the use of CT and FDG-PET uncovers most of these tumors at symptom presentation or within the first year of the neurologic disorder.3 Among all paraneoplastic disorders, the anti-Ma2 immune response is the most specific for limbic, diencephalic, or upper brainstem encephalitis.4 In young men this disorder associates with testicular tumors, while in older men or women other tumors are involved. Of 25 men with anti-Ma2-associated encephalitis younger than 50 years, 19 had germ-cell tumors (18 in testis). The current study focuses on the remaining 6 patients whose tumors were not found by multiple ancillary tests or even at initial evaluation of the orchiectomy specimen. Eventually all 6 patients were found to have a microscopic intratubular germ-cell neoplasm unclassified type (IGCNU), a common precursor of most testicular cancers that takes approximately 5 years to become invasive.5 Methods Tissues and antibodies The 6 patients were seen by the authors and form part of a series of 46 patients with anti-Ma2-associated encephalitis (appendix E-1 on the Web site at One of the 6 patients has been reported in detail.7 Patients or family members consented for antibody and tumor studies. These studies were approved Lu AF21934 by the University of Pennsylvania Institutional Review Board. Tumor tissue was available from all 6 patients: 1 frozen and 5 embedded in paraffin. Detection of anti-Ma1 and Ma2 antibodies was performed using immunoblot of recombinant proteins, as reported.8 Oct4 affinity-purified goat antibody was purchased from Santa Cruz Biotechnology (Santa Cruz, CA) and used to re-examine orchiectomy specimens for tumor cells. IgG containing anti-Ma2 antibodies isolated from patients’ sera and labeled with biotin was used to determine the expression of Ma2 in tumor cells.9 Immunohistochemistry Paraffin-embedded tissue was deparaffinized and the antigens retrieved, as reported.10 Tissue sections were serially Lu AF21934 incubated with 0.3% H2O2 for 20 minutes, 10% goat serum for 1 hour, biotinylated patients’ IgG (80 g/mL) or anti-Oct4 (1:200) overnight at 4 C. Sections incubated with anti-Oct4 were subsequently incubated with biotinylated horse anti-goat IgG (Vector, Burlingame, CA). For all sections the reactivity was developed with the avidin-biotin peroxidase method. Sections incubated with biotinylated normal human IgG or normal goat serum served as controls. Double immunolabeling with Ma2 and Oct4 antibodies was performed Lu AF21934 using the Lu AF21934 appropriate Alexa Fluor secondary antibodies (Molecular Probes, OR). Images were photographed under a Zeiss fluorescence microscope using Axiovision software. Results General clinical features The 6 patients developed short-term memory deficits due to limbic dysfunction, and additional symptoms that resulted from involvement of the hypothalamus-diencephalon, brainstem, pallidum (severe hypokinesis), or cerebellar pathways (mild, asymmetric ataxia) (table and appendix E-1). In 5 cases symptoms Rabbit Polyclonal to POFUT1 correlated with the MRI findings (figure 1) and in 1 the initial MRI was normal and no.