Summarymedium may produce false positive results (11). In this report, we describe two modifications of the original Bethesda assay which result Antibodies against factor VIII coagulant activity can appear in in markedly increased specificity and allowed us to diagnose and monihaemophiliacs who are treated with factor VIII preparations but also tor the presence of factor VIII :C inhibitors, spontaneously in non-haemophiliacs. The Bethesda assay is the most commonly used method to detect these antibodies, but it lacks specificity especially in the lower range resulting in unreliable data.Two modifications are proposed and tested to resolve the imper fections: 1. Buffering the normal plasma used in the assay-and control mixture with 0.1 M imidazole to pH 7.4. p 2. Replacing the imidazole buffer in the control mixture by immunodepleted factor VIII deficient plasma. These modifications allow better discrimination between positive and negative samples and improve reliability.

Based on our previous results indicating the presence of a tumor suppressor gene (TSG), chromosome 21 was analysed for loss of heterozygosity (LOH) in 18 patients with acute myeloid leukemia (17, AML-M0; one, AML-M1). Allelotyping at polymorphic loci was performed on purified material, allowing unequivocal detection of allelic loss and homozygous deletions. Six AML-M0 patients shared a common region of LOH harboring a single gene: RUNX1 (AML1), the most frequent site of translocations in acute leukemia and a well-known fusion oncogene. Fluorescence in situ hybridization allowed the identification of deletions with breakpoints within RUNX1 in two patients as the cause of LOH. In the four others the LOH pattern and the presence of two karyotypically normal chromosomes 21 were in line with mitotic recombination. Further molecular and cytogenetic analyses showed that this caused homozygosity of primary RUNX1 mutations: two point mutations, a partial deletion and, most significantly, a complete deletion of RUNX1. These findings identify RUNX1 as a classical TSG: both alleles are mutated or absent in cancer cells from four of the 17 AML-M0 patients examined. In contrast to AML-M0, the AML-M1 patient was trisomic for chromosome 21 and has two mutated and one normal RUNX1 allele, suggesting that the order of mutagenic events leading to leukemia may influence the predominant tumor type.

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A method is proposed to assess deterioration of frying fats by measuring polar and nonpolar components separated on a silica gel column. Means for polar components measured in duplicate samples by 19 collaborators ranged from 8.0 ± 0.34 to 25.8 ± 0.90%. Coefficients of variation ranged from 3.5 to 4.9%. The method has been adopted as official first action.

This paper describes the measurement of S phase DNA content in human bone marrow subpopulations using a single laser method for bivariate analysis of DNA content and cell‐surface immunofluorescence (s‐IF). Low density (<1.077 g/ml) bone marrow cells were labeled with a panel of unconjugated monoclonal antibodies (MoAb) for the lymphoid (CD2 + CD19), T‐lymphoid (CD2), B‐lymphoid (CD19), erythroid (anti‐glycophorin‐A), myelomonocytic (CD13, CD33; single and as cocktail) and monocytic (CD14) lineages. A fluorescein isothiocyanate (FITC)‐conjugated goat anti‐mouse label was used as second step. Unfixed, MoAb‐labeled cells were incubated for 24 h with a hypotonic propidium iodide solution for DNA staining. Cells were analysed on a single‐laser flow cytometer, operating at 488 nm. The effect of the combined staining protocol upon both s‐IF and DNA stainability was evaluated. Only a slight decrease (mean: 29.0%) in s‐IF intensity was observed after DNA staining. The percentages of immunofluorescent cells in the bone marrow samples of 10 normal individuals before and after DNA staining were essentially unchanged for all the MoAbs used. The DNA histograms of the immunophenotypically defined subpopulations were of excellent quality with a mean coefficient or variation of 1.8%. This procedure allows the assessment of very low levels of S‐phase DNA content, as measured in normal low density blood cells of 8 healthy volunteers (mean 0.07%). Considerable differences in percentages of cells with S‐phase content were seen between the four main bone marrow subpopulations (P<0.0001). The erythroid hematopoietic lineage showed the highest proportion of S‐phase DNA cells (26.4% ± 4.7%), the lymphoid lineage the lowest (2.5% ± 0.8%) while the monocytic lineage (6.1% ± 0.9%) and the myelomonocytic lineage (8.5% ± 1.4%) were in between. Within the lymphoid lineage the percentage of S‐phase cells was significantly lower in T‐cells (2.0% ± 0.7%) compared to B‐cells (5.1% ± 1.5%) (P<0.0001). We conclude that bivariate measurement of S‐phase DNA content and s‐IF is feasible with use of a single laser flow cytometer and that this approach provides important additional biological information. The value of this method to study hematologic malignancies is demonstrated in a bone marrow sample from a patient with erythroleukemia.