Specimen stability for DNA-based diagnostic testing.

The use of molecular diagnostic testing is increasing in the clinical setting; therefore, data regarding DNA stability in clinical specimens are essential for correct test performance and interpretation. This study was designed to determine DNA stability in peripheral blood and solid tissue under different storage conditions. DNA quality and yield were assayed by spectrophotometric absorbance, gel electrophoresis, and suitability for Southern hybridization and polymerase chain reaction (PCR), the most widely employed clinical DNA analyses.

Molecular Analysis in a Patient With Waldenström's Macroglobulinemia Reveals a Rare Case of Biclonality.

Background: The immunoglobulin and T-cell receptor gene rearrangement test is used to identify monoclonal populations in B- and T-cell malignancies and has become an important adjunct to morphologic analysis and immunophenotyping by flow cytometry. Waldenström's macroglobulinemia (WM) is typically a monoclonal proliferation of B cells with morphology of plasmacytoid lymphocytes and production of monoclonal IgM. Methods and Results: We report a case of WM with biclonal gammopathy (IgM kappa and IgM lambda) involving the blood and a diffuse lymphoplasmacytic infiltrate in the bone marrow in an 83-year-old man.

Use of myeloperoxidase mRNA as a marker for myeloid lineage in acute leukemias.

Background: Positivity for myeloperoxidase is considered the diagnostic hallmark of myeloid lineage and is the major criterion in the classification of acute leukemias. Early myeloid precursors, however, may be cytochemically negative for myeloperoxidase enzymatic activity or protein, but positive for myeloperoxidase mRNA.

Objective: To evaluate the expression of the myeloperoxidase gene in leukemic blasts at the mRNA level and correlate the expression with blast cytochemistry and immunophenotyping.

Nucleolar organizer regions (AgNORs) in intraepithelial neoplasia of the uterine cervix.

The authors have studied the AgNORs counts in intraepithelial lesions of the uterine cervix. The following values were obtained (mean +/- SD AgNORs/cell): normal specimens--2.11 +/- 0.

BCL-2 gene rearrangements in lymphoid malignancies.

Chromosomal translocations consistently associated with lymphoid malignancies result in gene rearrangements and activation of cellular oncogenes. The bcl-2 gene rearrangement is one of the most thoroughly studied and clinically useful in the diagnosis and monitoring of patients with lymphoma. The molecular analysis of this gene rearrangement has led to the discovery of the first member of a new class of oncogenes, the regulators of programmed cell death.

bcr gene rearrangement analysis in myeloproliferative disorders other than chronic myelogenous leukemia.

The bcr gene rearrangement resulting from the Philadelphia translocation is diagnostic of chronic myelogenous leukemia (CML) and is considered the hallmark of this myeloproliferative disorder (MPD) at the molecular level. The other MPDs, essential thrombocythemia (ET), polycythemia vera (PV), agnogenic myeloid metaplasia (AMM), and unclassified MPD, share morphologic features with CML, making the diagnosis difficult in cases with considerable morphologic overlap. In such cases, molecular analysis becomes essential for accurate diagnosis.

Discordant morphologic features in bone marrow involvement by malignant lymphomas: use of gene rearrangement patterns for diagnosis.

Discordant morphology between lymph node or extra-nodal site and bone marrow (BM) involvement by non-Hodgkin's malignant lymphoma (NHL) is a common occurrence, causing diagnostic difficulties. Additional diagnostic problems are posed by lymphoid aggregates commonly found in the BM of elderly patients, the age group with the highest incidence of lymphoma. Morphologic features are used to distinguish between benign and malignant lesions but no feature is diagnostic and exceptions are numerous.

Myeloperoxidase mRNA detection for lineage determination of leukemic blasts: retrospective analysis.

Myeloperoxidase (MPO) mRNA is an early myeloid marker; its detection in the morphologically and immunophenotypically primitive blasts of acute undifferentiated leukemia (AUL) establishes myeloid lineage and allows reclassification as acute myelogenous leukemia with minimal differentiation (AML-MO). We have previously reported a procedure for MPO mRNA detection by RT-PCR (reverse transcription-polymerase chain reaction) and an adaptation for use of routine hematology smears. This variant procedure allows retrospective analysis of mRNA and is used in the present study to evaluate the lineage of leukemic blasts in seven cases with morphology and cytochemistry consistent with AUL.

Separate clones in concomitant multiple myeloma and a second B-cell neoplasm demonstrated by molecular and immunophenotypic analysis.

The occurrence of multiple myeloma (MM) and a second B-cell neoplasm in the same patient is a rare event. We present 2 such patients, and provide evidence to support the presence of separate clones in these coexisting neoplasms. In the first case, MM became evident 14 months after the diagnosis of chronic lymphocytic leukemia (CLL).

Polymerase chain reaction in the diagnosis of chromosomal breakpoints.

Polymerase chain reaction-based methods for the detection of translocation-induced gene rearrangements are now widely used for diagnostics and patient monitoring. This article concentrates on two of the best studied chromosome translocations resulting in specific gene rearrangements and oncogene activation: the Philadelphia translocation of chronic myelogenous leukemia and acute leukemias, and the t(14;18) translocation of follicular lymphomas.

Molecular diagnostic testing for determination of myeloid lineage in acute leukemias.

Determination of myeloid vs. lymphoid cell lineage in acute leukemias is essential for diagnosis, prognosis, and treatment. However, some leukemic cells are too primitive to be identified based on conventional morphology, cytochemistry, and immunophenotype criteria.

Bone marrow biopsy imprint preparations: use for molecular diagnostics in leukemias.

Bone marrow (BM) biopsies occasionally fail to yield aspirate specimens; such "dry taps" pose diagnostic difficulties. In the absence of a BM aspirate, morphological evaluation and cytochemistry rely on core biopsy imprint preparations (IP) and other analyses, e.g.

Retrospective DNA analysis using fixed tissue specimens.

The recent explosion of scientific and technical knowledge in the field of molecular biology has allowed us to make important advances in our understanding of the molecular basis of many human diseases. This technology has now entered the clinical laboratory where identification of specific genetic sequences can aid in the precise diagnosis of hematologic and other malignancies, inherited diseases, specific infectious agents, and inherited predisposition to disease. In addition, it can be applied to prenatal diagnosis, paternity testing, identification of minimal residual disease following treatment, and assessment of drug sensitivity or resistance.

A new procedure for cell lineage determination in acute leukemias. Myeloperoxidase mRNA detection.

Determination of cell lineage in acute leukemias is essential for diagnosis and treatment. Detection of myeloperoxidase (MPO) mRNA establishes myeloid lineage of leukemic blasts that may be too primitive to be identified as myeloblasts based on morphology, cytochemistry, or immunophenotype. A highly specific and sensitive new procedure for MPO mRNA detection has been developed using HL-60 cells.

Amplification of intermediate-size DNA sequences from formalin and B-5 fixed tissue by polymerase chain reaction.

Retrospective analysis of DNA from paraffin-embedded fixed bone marrow biopsy specimens is possible if preceded by amplification of the DNA sequences of interest by the polymerase chain reaction (PCR). These fixed specimens yield degraded DNA that may not be suitable for direct analysis by conventional digestion and hybridization methods. This limitation is circumvented by PCR amplification and subsequent analysis of the amplified products.

Chronic granulocytic leukemia: reassessment of morphologic and cytogenetic characteristics in Ph1-positive and Ph1-negative cases.

33 cases of chronic granulocytic leukemia (CGL) were reassessed to determine if, by strict morphologic criteria. Philadelphia chromosome (Ph1)-negative CGL exists as a diagnostic entity and if Ph1-positive CGL could be distinguished from Ph1-negative CGL. Cases were reassessed using published criteria and, of 11 Ph1-negative cases, only 4 could be reclassified as myelodysplastic syndromes or undifferentiated chronic myeloproliferative disorder.

Digoxin-like immunoreactivity in serum from neonates and infants reduced by centrifugal ultrafiltration and fluorescence polarization immunoassay.

We evaluated the TDx Digoxin II (Abbott) modified procedure for interference from digoxin-like immunoreactive factors (DLIF) in pediatric patients. The effectiveness of centrifugal ultrafiltration as a means of removing DLIF interference from the serum of such patients was assessed. We used sera from 40 patients who had not received digoxin, whom we divided into two age groups: 30 neonates (less than 34 days postpartum) and 10 infants (younger than six months).

Polymerase chain reaction: amplification of DNA from fixed tissue.

The polymerase chain reaction (PCR) allows the analysis of DNA from biologic samples containing only nanogram quantities of DNA. We used DNA purified from fresh or frozen peripheral blood (PB) leukocytes and formalin, or B-5 fixed bone marrow aspirate clots (BM). A sequence of the beta-globin gene was amplified via the PCR then hybridized with allele specific oligonucleotide probes for hemoglobin A, S, and C.

The influence of novel platelet aggregation inhibitors on human blood platelet clustering and retention effected by some polymer materials, in vitro.

Human blood platelet clustering and retention in vitro induced by medical grade Cuprophane and Silastic and by non-medical grade polyethylene and Mylar, are discussed. The influence of synthetic compounds on polymer-induced platelet clustering and retention has been evaluated, interpreted in terms of physicochemical parameters, and correlated with ADP- and human alpha-trombin-stimulated aggregation; the Pearson product-moment correlation coefficient for a series of bis (alkylnipecotoylamino) alkanes was computed at r = 0.98 between (i) mylar-induced cluster formation and (ii) ADP-effected aggregation, and at r = 0.