Photo-Induced Charge State Dynamics of the Neutral and Negatively Charged Silicon Vacancy Centers in Room-Temperature Diamond.

The silicon vacancy (SiV) center in diamond is drawing much attention due to its optical and spin properties, attractive for quantum information processing and sensing. Comparatively little is known, however, about the dynamics governing SiV charge state interconversion mainly due to challenges associated with generating, stabilizing, and characterizing all possible charge states, particularly at room temperature. Here, multi-color confocal microscopy and density functional theory are used to examine photo-induced SiV recombination - from neutral, to single-, to double-negatively charged - over a broad spectral window in chemical-vapor-deposition (CVD) diamond under ambient conditions. For the SiV0 to SiV- transition, a linear growth of the photo-recombination rate with laser power at all observed wavelengths is found, a hallmark of single photon dynamics. Laser excitation of SiV‒, on the other hand, yields only fractional recombination into SiV2‒, a finding that is interpreted in terms of a photo-activated electron tunneling process from proximal nitrogen atoms.

Randomised controlled trial of paediatric magnetic positioning device assisted colonoscopy: a pilot and feasibility study.

BACKGROUND AND AIMS: Complete colonoscopy is critical for the evaluation of many paediatric gastrointestinal diseases. The aim of the study was to investigate the feasibility of magnetic positioning device for paediatric colonoscopy and to compare completion rate and procedure time with and without the device. METHODS: Prospective randomised controlled trial of standard colonoscopy compared to magnetic positioning device assisted colonoscopy in children and adolescents ages 7-20 years was performed. RESULTS: Analysis showed that the proportion of successfully completed colonoscopies were 19/20 (95%) in the MP arm versus 17/18 (94.4%) in the SC arm, p=NS. The median time to complete colonoscopy to the cecum was 16.5 min (range 6-52 min) in the MP arm and 12 min (range 6-33 min) in the SC arm, p=NS. CONCLUSIONS: Our preliminary data suggest that the use of magnetic positioning device for colonoscopy is feasible in paediatric patients. These data suggest that the use of magnetic positioning device may not be of benefit for experienced endoscopists who achieved very high colonoscopy completion rates without the MP device. Further studies are needed to determine its role in paediatric colonoscopy since this device may be of more benefit for physicians in training.

A case of aggressive multiple myeloma with cleaved, multilobated, and monocytoid nuclei, and no serum monoclonal gammopathy.

Multiple myeloma is a B-cell malignancy characterized by proliferation of neoplastic plasma cells. A few cases have been reported identifying variant forms of neoplastic plasma cells with atypical nuclei that secrete myeloma protein. We report a highly unusual case of plasma cell myeloma that presented with cleaved, multilobated, and monocytoid nuclei, without detectable myeloma protein in the serum or urine. The bone marrow contained sheets of plasma cells exhibiting pleomorphic nuclei with cleaved, multilobated, and monocytoid features that were negative for myeloperoxidase and dual esterase. Flow cytometric analysis revealed CD38high/CD45low cells expressing cytoplasmic kappa light chain, without evidence of myeloid or lymphoid differentiation. Following chemotherapy, the patient developed secondary plasma cell leukemia. A high plasma cell labeling index was obtained from bone marrow and peripheral blood, indicating a poor prognosis. In addition to quantitative immunoglobulins, serum protein electrophoresis, and immunofixation electrophoresis of serum and urine, we recommend cytochemical and flow cytometric studies for evaluation of suspected plasma cell myeloma with atypical cellular features.

Acute exposure to the abused inhalant, isobutyl nitrite, reduced T cell responsiveness and spleen cellularity.

Isobutyl nitrite is an inhalant abused principally by male homosexuals. We have reported that subchronic inhalation exposure (45 min/day for 14 days) to 900 ppm isobutyl nitrite was immunosuppressive. In the present study, the effects of acute exposure to the inhalant were examined. Mice were exposed in an inhalation chamber to 900 ppm isobutyl nitrite for 45 min. One day later, spleen cellularity was reduced by 39% without selectively depleting CD4(+) or CD8(+) cells. The numbers of peripheral blood leukocytes and peritoneal cells were also reduced. Following acute inhalation exposure, T cell proliferative responses stimulated with allogeneic cells or anti-CD3 and anti-CD28 antibodies were inhibited, while mitogen-induced responses were not affected. Purified T cells exposed to the inhalant also had compromised responses, suggesting a direct effect on T cells. However, the cumulative effects of multiple exposures were necessary to inhibit T-dependent antibody responses or T cell or macrophage cytotoxicity.

Nitrite inhalants spontaneously liberate nitric oxide, which is not responsible for the immunotoxicity in C57BL/6 mice.

Nitrite inhalant abuse has been correlated epidemiologically with HIV seropositivity and with Kaposi's sarcoma. Using a mouse model, we have shown that inhaled isobutyl nitrite caused anemia and severely depressed immunity. In the present study, we showed that both isobutyl and cyclohexyl nitrites in air liberated nitric oxide (NO). An immunotoxic dose of 900 ppm isobutyl nitrite liberated 115 ppm NO. Mice were exposed in an inhalation chamber to 115 ppm NO, 900 ppm isobutyl nitrite, or 900 ppm cyclohexyl nitrite for 45 min/day. Following a single exposure, NO did not affect peripheral blood cell counts, while isobutyl and cyclohexyl nitrites reduced cell numbers. After 14 daily exposures, isobutyl nitrite, but not cyclohexyl nitrite or NO, reduced peritoneal macrophage tumoricidal activity. The nitrite esters likely caused immunotoxicity by mechanisms other than NO release.

Severe hemorrhagic complication due to acquired factor V inhibitor after single exposure to bovine thrombin product.

Hemorrhagic complications have been reported after repeated exposures to bovine thrombin products due to development of factor V inhibitors. Our patient underwent emergency repair of acute aortic dissection and coronary artery bypass grafting. The patient developed leg wound infection at the saphenous vein harvest site, which was debrided and left open. Attempt to reclose the leg wound 1 month later was complicated by a life-threatening hemorrhage with markedly elevated activated partial thromboplastin time. There was no evidence of infection or disseminated intravascular coagulation, and further study identified low factor V level with positive factor V inhibitor. Treatment with plasmapheresis and steroid successfully reversed the coagulopathy. Detailed case review failed to reveal exposure to any thrombin products other than the one used for the aortic dissection repair. This case was unusual because only a single exposure to this product resulted in severe hemorrhagic complication 1 month after surgery.

An essential function of a phosphoinositide-specific phospholipase C is relieved by inhibition of a cyclin-dependent protein kinase in the yeast Saccharomyces cerevisiae.

The PLC1 gene product of Saccharomyces cerevisiae is a homolog of the delta isoform of mammalian phosphoinositide-specific phospholipase C (PI-PLC). We found that two genes (SPL1 and SPL2), when overexpressed, can bypass the temperature-sensitive growth defect of a plc1delta cell. SPL1 is identical to the PHO81 gene, which encodes an inhibitor of a cyclin (Pho80p)-dependent protein kinase (Pho85p) complex (Cdk). In addition to overproduction of Pho81p, two other conditions that inactivate this Cdk, a cyclin (pho80delta) mutation and growth on low-phosphate medium, also permitted growth of plc1delta cells at the restrictive temperature. Suppression of the temperature sensitivity of plc1delta cells by pho80delta does not depend upon the Pho4p transcriptional regulator, the only known substrate of the Pho80p/Pho85p Cdk. The second suppressor, SPL2, encodes a small (17-kD) protein that bears similarity to the ankyrin repeat regions present in Pho81p and in other known Cdk inhibitors. Both pho81delta and spl2delta show a synthetic phenotype in combination with plc1delta. Unlike single mutants, plc1delta pho81delta and plc1delta spl2delta double mutants were unable to grow on synthetic complete medium, but were able to grow on rich medium.

Adeno-associated virus Rep78 inhibits oncogenic transformation of primary human keratinocytes by a human papillomavirus type 16-ras chimeric.

Seroepidemiologic studies demonstrate that adeno-associated virus (AAV) infection is negatively associated with cervical cancer. This inverse association may be due to an ability of AAV to inhibit the role of human papillomavirus (HPV) in cervical carcinogenesis. In support of this hypothesis AAV has been demonstrated to inhibit several papillomavirus types, including bovine papillomavirus type 1 and human papillomaviruses types 16 and 18 (HPV-16/18) in tissue culture. The AAV-encoded Rep78 protein was responsible for this inhibition. These previous studies, however, were largely carried out in immortalized mouse fibroblasts. This cell type is likely not to be the most accurate model cell type for studying HPV-associated cervical carcinogenesis. In this study it is demonstrated that AAV Rep78 protein inhibits oncogenic transformation of freshly explanted primary human foreskin keratinocytes by an HPV-16/ras chimeric genome. Such cells are the natural host cell type for HPV-16/18 infection. It is also demonstrated that the HPV-16 P97 promoter is specifically inhibited by Rep78 in a transient CAT assay. These data further extend our knowledge of the AAV-papillomavirus interaction and provide a model for investigating the negative association of AAV with cervical cancer.

Acute blood toxicity of the abused inhalant, cyclohexyl nitrite.

Cyclohexyl nitrite is an abused nitrite inhalant. This is the first report of toxicity of cyclohexyl nitrite. Mice were exposed to 300-900 ppm cyclohexyl nitrite in an inhalation chamber for 45 min and then bled. Such treatment resulted in a 7-10% reduction in red blood cell counts, haemoglobin and haematocrit levels. Both blood leucocyte counts and spleen cellularity were reduced by 40%. Unlike isobutyl nitrite, subchronic treatment of mice with cyclohexyl nitrite did not impair macrophage tumoricidal activity or production of reactive nitrogen intermediates, but did modulate B and T cell mitogen responses. The data suggest that cyclohexyl nitrite had cytotoxic activity, comparable to that of isobutyl nitrite, which might be related to the anaemia reported in abusers. The immunomodulatory properties of cyclohexyl nitrite differed from those of isobutyl nitrite.

Quantification of vitamin D receptor mRNA by competitive polymerase chain reaction in PBMC: lack of correspondence with common allelic variants.

It has been recently claimed that polymorphism for the vitamin D receptor (VDR) influences several aspects of calcium and bone metabolism. To evaluate the physiologic plausibility of these claims, we compared the abundance of the VDR mRNA in peripheral blood mononuclear cells (PBMCs) between different VDR genotypes using a quantitative reverse transcribed polymerase chain reaction-based method. The method is based on the coamplification of VDR cDNA and an internal standard consisting of known concentrations of a human VDR CDNA mutated at a BglII restriction site; the interassay coefficient of variation is 11%. To validate the method, we made use of earlier receptor binding studies indicating that normal human monocytes and activated, but not resting, lymphocytes expressed the VDR. The concentration of the VDR mRNA was 10(-8) to 10(-7) g/g of total RNA in cell-sorted monocytes and in in vitro activated lymphocytes, but only 10(-12) g/g of total mRNA in resting lymphocytes, establishing that the VDR mRNA determined by our method in PBMCs is due to constitutive expression in monocytes. Following an initial genotype screening of 85 normal volunteers by polymerase chain reaction or restriction fragment length polymorphism analysis, 14 individuals with the Bb genotype, 12 with the bb genotype, and 12 with the BB genotype were selected. The concentration of the VDR mRNA, corrected for the number of monocytes, was similar among the three genotype groups, as were the other variables examined: serum calcitriol, serum osteocalcin, and vertebral and hip bone density. We conclude that VDR polymorphism does not affect the abundance of the VDR mRNA.

Expression of the SUC2 gene of Saccharomyces cerevisiae is induced by low levels of glucose.

High levels of glucose repress expression of the SUC2 gene in the yeast Saccharomyces cerevisiae. We have discovered that low levels of glucose are required for maximal transcription of SUC2: SUC2 expression is induced about five- to ten-fold in cells growing on low levels of glucose (0.1%) compared to cells growing on galactose or glycerol. Two pieces of evidence suggest that this low-glucose-induced expression is mediated by a repression mechanism that involves an upstream repression site in the SUC2 promoter (URS(SUC2)). First, deletion of the URS(SUC2) results in expression of the SUC2 gene in the absence of glucose, and second the URS(SUC2) mediates a six-fold repression of a reporter gene when inserted into a heterologous promoter. However, this URS(SUC2) mediated repression occurs on all tested carbon sources, suggesting that this URS element acts in concert with all other promoter elements to respond to low concentrations of glucose. This repression requires the general repressor SSn6p. SNF3, which encodes a glucose transporter that appears to be a sensor of low levels of glucose, is also required for low-glucose-induced expression of SUC2.

Fibrosing colonopathy in children with cystic fibrosis.

PURPOSE: Fibrosing colonopathy is a newly described entity seen in children with cystic fibrosis. The radiological hallmarks are foreshortening of the right colon with varying degrees of stricture formation. High-dose enzyme therapy has been implicated as the cause of this process. The purpose of this study is to review the author's experience with evaluation and treatment of these patients. METHODS: There are currently 380 patients being treated at our CF center. Fifty-five of these patients have been treated with high-dose enzyme therapy (> 5,000 units of lipase/kg). The medical records of these patients, who are at risk for developing fibrosing colonopathy, were reviewed for the presence of recurrent abdominal complaints, and the work-up and treatment of these symptoms. RESULTS: Chronic complaints of abdominal pain, distension, change in bowel habits, or failure to thrive were present in 24 of the 55 patients treated with high-dose enzymes. So far, 18 of these 24 patients have been evaluated by contrast enema. Thirteen of eighteen have been found to have fibrosing colonopathy characterized by foreshortening and strictures of the colon. Additional findings included focal strictures of the right colon (7 of 13), long segment strictures (5 of 13), and total colonic involvement (1 of 13). Nine patients with the most severe symptoms have undergone colon resection, including five segmental right colectomies, three extended colectomies (ileo-sigmoid anastomosis), and one subtotal colectomy with end-ileostomy. Pathological evaluation has shown submucosal fibrosis, destruction of the muscularis mucosa, and eosinophilia. No postoperative complications or deaths occurred. All nine postoperative patients have noted marked symptomatic improvement. Contrast enema follow-up results are available for six patients, and have documented no recurrent strictures to date. Three of four nonoperative patients have less severe symptoms and are currently being treated conservatively. The other family has refused surgery and the patient is being treated symptomatically. CONCLUSION: High-dose lipase replacement has been implicated as the etiology for FC and was present in all of our patients. Our cystic fibrosis center now routinely limits lipase to 2,500 U/kg per dose. We recommend the use of the contrast enemas to evaluate at-risk patients who have chronic abdominal complaints or who present with recurrent bowel obstruction. Colon resection should be performed in those with clinically and radiographically significant strictures with the expectation of a good outcome.

Preceding standard therapy is the likely cause of MDS after autotransplants for multiple myeloma.

Myelodysplastic syndromes (MDS) and acute myeloid leukaemia (AML) have been reported after autologous transplantation (AT) for lymphoma. It is not clear whether myeloablative therapy used in conjunction with autologous transplantation contributes to the development of MDS/AML or whether the conventional chemotherapy preceding the transplant, and administered over a prolonged period, causes these secondary malignancies. To address this issue, we examined 188 patients with multiple myeloma (MM) who had received AT. 71 patients with no more than one cycle of standard chemotherapy were enrolled in our Total Therapy program, designed to avoid exposure to alkylating agents prior to peripheral blood stem cell mobilization (group 1). The median duration of pretransplant therapy in group 1 was 7.6 months and significantly shorter than the 24 months of 117 patients (group 2) with more prolonged conventional therapy (P = 0.0001). All seven patients developing MDS post-transplantation belonged to group 2 (P = 0.02); the median durations from initial therapy and first transplant were 66 months (range 38-86) and 24 months (range 9-39), respectively. Our findings provide evidence that prolonged standard-dose alkylating agent therapy prior to transplantation, rather than autotransplant-supported myeloablative treatment, is associated with development of MDS/AML. Stem cell damaging alkylator treatment should be avoided, not to compromise PBSC collection, but also to reduce the risk of treatment-related MDS/AML.

Leukopenia and altered hematopoietic activity in mice exposed to the abused inhalant, isobutyl nitrite.

Isobutyl nitrite is representative of a group of inhalants abused primarily by male homosexuals; abuse of this drug may be a risk factor for AIDS or Kaposi's sarcoma. Using a 14-day exposure regimen, we previously reported that inhaled isobutyl nitrite was immunotoxic to mice, severely compromising T-dependent antibody responses and cytotoxic T cell and macrophage tumoricidal activity. In addition, exposure to the inhalant dramatically reduced spleen cellularity. A single 45-minute inhalation exposure produced anemia in mice. In the present study, we examined the effects of subchronic exposure to the drug on peripheral blood cellularity and hematopoietic activity. Mice were exposed to 900 ppm isobutyl nitrite in an inhalation chamber for 45 minutes/day for 14 days. One day after the final exposure, the number of peripheral blood leukocytes was reduced by 32%; however, the number of erythrocytes was increased by 7%. This was accompanied by an apparent shift from myelopoiesis to erythropoiesis. The numbers of bone marrow and spleen burst-forming units-erythroid (BFU-E) were increased about two-fold, while the numbers of colony-forming units-granulocyte/macrophage (CFU-GM) were decreased by about half. Bone marrow stromal cells also had reductions in the production of myeloid colony-stimulating activity after subchronic exposure to the inhalant. In addition, the numbers of hematopoietic stem cells, colony-forming units-spleen (CFU-S), were reduced in both bone marrow and spleen. Peripheral blood erythrocyte and leukocyte counts returned to normal levels by 7 days after the final exposure, as did the number of BFU-E. The number of CFU-GM remained depressed, however, even after 7 days of recovery. These data suggest that repeated exposures nonspecifically depleted cells and that erythropoiesis was stimulated, apparently at the expense of myelopoiesis.

Acute inhalation exposure to isobutyl nitrite causes nonspecific blood cell destruction.

Abuse of nitrite inhalants is widespread among male homosexuals and has been epidemiologically correlated with seropositivity to human immunodeficiency virus (HIV) and to Kaposi's sarcoma. These drugs may act as cofactors in AIDS if they compromise the ability to resist infection or tumor growth. We have previously reported that 14 daily 45-minute exposure to 900 ppm isobutyl nitrite in an inhalation chamber did compromise the immunocompetence of mice. We now report that a single 45-minute exposure produced a transient anemia. Erythrocyte counts, hemoglobin, and hematocrit levels were reduced by 7% but rebounded to above-normal levels 24 hours later. In vitro exposure of blood to isobutyl nitrite vapors did not lyse the cells but did induce Heinz body formation and increase their binding to macrophages. Thus, it is likely that the red cells were removed by phagocytic clearance not by direct lysis. Blood leukocyte numbers were also reduced following a single exposure to the inhalant, but the cell loss was delayed until 24 hours after exposure. Recovery of peripheral blood leukocytes 72 hours after exposure coincided with a reduction in spleen cellularity, suggesting that spleen cells were mobilized to replace lost blood leukocytes.

Regulation of tight junction resistance in T84 monolayers by elevation in intracellular Ca2+: a protein kinase C effect.

Elevation in intracellular Ca2+ acting via protein kinase C (PKC) is shown to regulate tight junction resistance in T84 cells, a human colon cancer line and a model Cl- secretory epithelial cell. The Ca2+ ionophore A23187, which was used to increase the intracellular Ca2+ concentration, caused a decrease in tight junction resistance in a concentration- and time-dependent manner. Dual Na+/mannitol serosal-to-mucosal flux analysis performed across the T84 monolayers treated with 2 microM A23187 revealed that A23187 increased both fluxes and that in the presence of ionophore there was a linear relationship between the Na+ and mannitol fluxes with a slope of 56.4, indicating that the decrease in transepithelial resistance was due to a decrease in tight junction resistance. Whereas there was no effect of 0.1 microM A23187, 1 or 2 microM produced a 55% decrease in baseline resistance in 1 hr and 10 microM decreased resistance more than 80%. The A23187-induced decrease in tight junction resistance was partially reversible by washing 3 times with a Ringer's-HCO3 solution containing 1% BSA. The A23187 effect on resistance was dependent on intracellular Ca2+; loading the T84 cells with the intracellular Ca2+ chelator BAPTA significantly reduced the decrease in tight junction resistance caused by A23187. This intracellular Ca2+ effect was mediated by protein kinase C and not calmodulin. While the protein kinase C antagonist H-7 totally prevented the action of A23187 on tight junction resistance, the Ca2+/calmodulin inhibitor W13 did not have any effect. Sphingosine, another inhibitor of PKC, partially reduced the A23187-induced decline in tight junction resistance. The PKC agonist PMA mimicked the A23187 effect on resistance, although the effect was delayed up to 1 hr after exposure. In addition, however, PMA also caused an earlier increase in resistance, indicating it had an additional effect in addition to mimicking the effect of elevating Ca2+. The effects of a phospholipase inhibitor (mepacrine) and of inhibitors of arachidonic acid metabolism (indomethacin for the cyclooxygenase pathway, NDGA for the lipoxygenase pathway, and SKF 525A for the epoxygenase pathway) on the A23187 action were also examined. None of these agents altered the A23187-induced decrease in resistance. Monolayers exposed to 2 microM A23187 for 1 hr were stained with fluorescein conjugated phalloidin, revealing that neighboring cells did not part one from another and that A23187 did not have a detectable effect on distribution of F-actin in the perijunctional actomyosin ring. The results indicate that elevation in intracellular Ca2+ decreases tight junction resistance in the T84 monolayer, acting through protein kinase C by a mechanism which does not involve visible changes in the perijunctional actomyosin ring.

Autoregulatory circuits in myeloma. Tumor cell cytotoxicity mediated by soluble CD16.

BACKGROUND: Multiple myeloma remains an incurable malignancy due to marked resistance of the tumor to standard doses of chemotherapy. Treatment approaches, using chemotherapeutic dose escalation and hematopoietic stem cell support have resulted in significant augmentation of tumor mass reduction such that complete remissions are effected in approximately 50% of patients. These remissions are however, often not durable. In the setting of minimal residual disease, therefore, adjunctive immunotherapy may be useful. METHODS: Peripheral blood mononuclear cells were studied from 28 untreated patients with multiple myeloma (MM). Mononuclear cell CD16 (FcR gamma III) expression was determined by flow cytometry. The effect of lymphocyte-derived soluble CD16, isolated by affinity chromatography, on MM cell growth and differentiation was assessed. MM cell proliferation, viability, immunoglobulin production and gene expression was studied. RESULTS: Data are presented indicating that cells expressing CD16 are increased in untreated patients with IgG-secreting myeloma. The predominant phenotype of these cells is CD8+ or CD56+. These CD16+ cells can produce a soluble form of the Fc receptor (sFcR, sCD16) that can bind to surface Ig on cultured human IgG-secreting myeloma cells and effect suppression of tumor cell growth and Ig secretion. This effector function is accompanied by concomitant suppression of c-myc as well as IgH and IgL gene transcription. Finally, prolonged exposure to sCD16 causes myeloma tumor cell cytolysis. CONCLUSIONS: sCD16 and possibly other soluble FcR are candidate molecules for adjunctive immunotherapy of myeloma, once complete responses have been effected by intensive cytotoxic therapy, now possible in up to 50% of newly diagnosed patients.

Multiple mechanisms provide rapid and stringent glucose repression of GAL gene expression in Saccharomyces cerevisiae.

Expression of the GAL genes of Saccharomyces cerevisiae is induced during growth on galactose by a well-characterized regulatory mechanism that relieves Gal80p inhibition of the Gal4p transcriptional activator. Growth on glucose overrides induction by galactose. Glucose repression acts at three levels to reduce GAL1 expression: (i) it reduces the level of functional inducer in the cell; (ii) it lowers cellular levels of Gal4p by repressing GAL4 transcription; and (iii) it inhibits Gal4p function through a repression element in the GAL1 promoter. We quantified the amount of repression provided by each mechanism by assaying strains with none, one, two, or all three of the repression mechanisms intact. In a strain lacking all three repression mechanisms, there was almost no glucose repression of GAL1 expression, suggesting that these are the major, possibly the only, mechanisms of glucose repression acting upon the GAL genes. The mechanism of repression that acts to reduce Gal4p levels in the cell is established slowly (hours after glucose addition), probably because Gal4p is stable. By contrast, the repression acting through the upstream repression sequence element in the GAL1 promoter is established rapidly (within minutes of glucose addition). Thus, these three mechanisms of repression collaborate to repress GAL1 expression rapidly and stringently. The Mig1p repressor is responsible for most (possibly all) of these repression mechanisms. We show that for GAL1 expression, mig1 mutations are epistatic to snf1 mutations, indicating that Mig1p acts after the Snf1p protein kinase in the glucose repression pathway, which suggests that Snf1p is an inhibitor of Mig1p.

An introductory orientation to clinical pathology core and on-call responsibilities.

An introductory 4-week orientation for clinical pathology is described. There were 76 hours of lectures, 74 hours of conferences, and 68 hours of laboratories for a total of 221 hours. During the orientation, all calls handled by the residents were evaluated as to resolution, patient outcome, and interaction required. Eighty calls were received during the orientation from 57 technologists (71%), 16 physicians (20%), and seven nurses (9%). The calls originated concerning the following: blood banking, 37 (46%); hematology, 21 (27%); chemistry, 14 (18%); microbiology, five (6%); and administration, three (4%). Sixty percent of the calls were consultative and 40% were supervisory. Ninety-nine percent were handled appropriately by the residents. Patient outcome was moderately or significantly affected in 44% of all calls, divided between 67% of all consultative calls and 9% of all supervisory calls. Significant pathologist interaction was required in 49% of all calls, divided between 71% of the consultative calls and 16% of the supervisory calls. Using this integrated, dynamic system of resident instruction, on-call experience, and evaluation, residents quickly gain confidence in handling call, didactic clinical consultation, and patient management. The orientation and on-call system described provides for a relevant and dynamic system for resident education.