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Aflatoxin produced by Aspergillus flavus is known to be strongly related to liver injury (hepatocellular carcinoma) and immune system damage involving leukocytes. This toxin suppresses both the cell-mediated immune system and macrophage function, and decreases the production of complement and interferon molecules. Purpose To evaluate the presence of aflatoxin in infectious lesions as well as how the toxin is taken up by leukocytes. Method Pathological specimens from a patient who died from aspergillosis caused by aflatoxin-producing A. flavus were used. Anti-aflatoxin B1 antibody was reacted with paraffin-embedded lesion specimens from the heart, kidney, and thyroid gland of the patient and observed microscopically. Result Positive reactions were detected in fungal elements and leukocytes (neutrophils and macrophages) in inflammatory lesions. Conclusion Within the patient's body, A. flavus likely produced aflatoxin, which then was taken up by neutrophils and macrophages.These results suggest that leukocyte function and the immune mechanism are locally suppressed by aflatoxin.

Aflatoxin produced by Aspergillus flavus is known to be strongly related to liver injury (hepatocellular carcinoma) and immune system damage involving leukocytes. This toxin suppresses both the cell-mediated immune system and macrophage function, and decreases the production of complement and interferon molecules.To evaluate the presence of aflatoxin in infectious lesions as well as how the toxin is taken up by leukocytes.Pathological specimens from a patient who died from aspergillosis caused by aflatoxin-producing A. flavus were used. Anti-aflatoxin BPositive reactions were detected in fungal elements and leukocytes (neutrophils and macrophages) in inflammatory lesions.Within the patient's body, A. flavus likely produced aflatoxin, which then was taken up by neutrophils and macrophages.These results suggest that leukocyte function and the immune mechanism are locally suppressed by aflatoxin.

Vitamin B6 (VB6) deficiency contributes to oncogenesis and tumor progression in certain cancers, and is prevalent in cancer patients in general. VB6 is also an essential element of heme synthesis, and deficiency can lead to anemia. Primary myelofibrosis (PMF) and secondary myelofibrosis (sMF) are myeloproliferative neoplasms often presenting with anemia along with other cytopenias. We performed a prospective study to determine whether PMF and sMF patients suffer from VB6 deficiency, and whether VB6-deficient patients show improvement of anemias with VB6 supplementation. Twelve PMF patients and 11 sMF patients were analyzed. A total of 16 of 23 patients (69.6%) were found to have VB6 deficiency, but VB6 supplementation with pyridoxal phosphate hydrate did not elevate hemoglobin levels in deficient patients. None of the patients presented with vitamin B12, iron, or copper deficiencies. Four patients showed serum folate levels below the lower limit of normal and eight patients showed serum zinc levels below the lower limit of normal; however, these deficiencies were marginal and unlikely to contribute to anemia. Compared to VB6-sufficient patients, VB6-deficient patients showed significantly lower serum folate levels and higher serum copper levels. Studies elucidating the relationship of VB6 deficiency and etiology of PMF/sMF are warranted.

Autologous peripheral blood stem cell transplantation(auto-PBSCT)combined with high-dose chemotherapy has been considered as the standard therapy for relapsed or induction therapy-refractory aggressive lymphomas sensitive to chemotherapy. While various regimens have been applied as the conditioning,none has yet been established as the standard. We have begun to employ high-dose ranimustine,cytarabine,etoposide and cyclophosphamide(MCVAC)regimen. The present study was undertaken to review the efficacy and safety of MCVAC. Regimen: We carried out a retrospective analysis of 20 patients diagnosed as diffuse large B-cell lymphoma. The median follow-up duration of 20 patients was 13.05 months(range, 0.57-49.5 months). The 4-year OS and PFS were 57.8% and 30.2%,respectively. Relapse was the most frequent cause of treatment failure(n=7). The major toxicities were anorexia/nausea(95%),diarrhea (75%),hypokalemia (70%). One patient died of hepatic veno-occlusive disease(VOD). The serious adverse events included hypokalemia,arrhythmia,cerebral hemorrhage,and heart failure(1 case[5%]each). There was 1 case of a late-onset adverse event: therapy-related myelo- dysplastic syndrome/acute myeloblastic leukemia(MDS/AML). MCVAC regimen was concluded as effective and well-toler- ated. However,we should carefully monitored for the possible development of VOD and MDS/AML. Further follow-up is needed to evaluate the long-term efficacy and safety.

Neurological symptoms induced by the infiltration of malignant lymphoma into the nervous systems are subsumed under the term neurolymphomatosis (NL). Here, we report the case of a 30-year-old Japanese man with primary testicular lymphoma complicated, as seen in various neurological findings, by secondary NL prior to testicular swelling. Painless right scrotal enlargement was noticed more than 1 month after the appearance of neurological complications such as right upper extremity numbness, dysarthria, facial palsy, and diplopia. Proactive investigation and biopsies of extranodal sites at high risk of central nervous system infiltration of malignant lymphoma, such as the testes, should be considered when secondary NL is suspected based on imaging findings.

Cardiac amyloid light-chain amyloidosis (AL amyloidosis) is a rare disease with a very poor prognosis, associated with plasma cell dyscrasias such as monoclonal gammopathy of undetermined significance and multiple myeloma. Though bortezomib-containing regimens have achieved high hematologic response rates, there are still few reports describing the outcomes of Japanese patients. Six patients with severe cardiac AL amyloidosis were treated with bortezomib-containing regimens. Involved free light chain (iFLC) decreased immediately in most of these cases. However, the condition of heart failure and N-terminal pro-B-type natriuretic peptide (NT-proBNP) worsened in the early phase of this treatment and then improved several months later. At 29 months, the median duration of follow-up (2-47months), all patients remain alive except one who died of sudden cardiac arrest. Bortezomib-containing regimens are considered to be among the effective treatments for severe cardiac AL amyloidosis.

There are some reports regarding hepatitis B virus (HBV) reactivation in patients with myeloma who are HBV carriers or who have had a resolved HBV infection, and there is no standard prophylaxis strategy for these patients. We performed a retrospective multicenter study to determine the incidence and characteristics of HBV reactivation in patients with multiple myeloma. We identified 641 patients with multiple myeloma who had been treated using novel agents and/or autologous stem cell transplantation with high-dose chemotherapy between January 2006 and June 2014 at nine Japanese hospitals. The patients’ characteristics, laboratory data, and clinical courses were retrieved and statistically analyzed. During a median follow-up of 101 weeks, one of eight (12.5 %) HBV carriers developed hepatitis and 9 of 99 (9.1 %) patients with resolved HBV infection experienced HBV reactivation; the cumulative incidences of HBV reactivation at 2 years (104 weeks) and 5 years (260 weeks) were 8 and 14 %, respectively. The nine cases of reactivation after resolved HBV infection had received entecavir as preemptive therapy or were carefully observed by monitoring their HBV DNA levels, and none of these cases developed hepatitis. Among patients with multiple myeloma, HBV reactivation was not rare. Therefore, long-term monitoring of HBV DNA levels is needed to prevent hepatitis that is related to HBV reactivation in these patients.

Total body irradiation (TBI) is an important component of conditioning regimens for Allogeneic bone marrow transplantation (BMT). Interstitial pneumonitis (IP) and other pulmonary disorders are known regimen-related complications. The incidence of IP is related to the dose rate and dose fractionation; however, there is a paucity of clinical data regarding the optimal dose fractionation. This retrospective study evaluated patients to determine the influence of dose fractionation during TBI in preparation for allogeneic BMT on the subsequent development of IP and other pulmonary complications. Fifty-six patients were treated with TBI followed by BMT at our institute. All patients received a total TBI dose of 12 Gy given in 6 fractions over 3 days or in 4 fractions over 2 days. The prevalence of unrelated donors in the 4-fraction group was higher than that in the 6-fraction group. The overall and freedom from progression rates for patients in the 4-fraction group were better than those for patients in the 6-fraction group, but the difference did not reach significance. Clinically significant lung complications occurred in 19 (10: infectious and 9: non-infectious diseases) of 33 patients in the 6-fraction group and 12 (7: infectious and 5: non-infectious diseases) of 23 in the 4-fraction group. There was no significant difference between the two groups. There was no significant difference in pulmonary complications between patients treated with a TBI dose of 12 Gy in 6 fractions over 3 days and patients treated with a TBI dose of 12 Gy in 4 fractions over 2 days.

JAK2V617F, a gain-of-function mutation in the tyrosine kinase JAK2, is frequently detected in classical myeloproliferative neoplasms (MPNs). In the present study, we determined the JAK2V617F allele burden in Japanese MPN patients using alternately binding probe competitive-polymerase chain reaction, a highly quantitative method recently developed by our group. Although we observed strong similarities in terms of epidemiological parameters associated with the JAK2V617F allele burden between our cohort and others, we found a higher JAK2V617F allele burden in Japanese polycythemia vera (PV) patients and lower frequencies of thrombosis in Japanese MPN patients compared with previous reports. In addition, despite the presence of high red blood cell counts, some patients bearing the JAK2V617F mutation were not diagnosed as PV, as their hemoglobin values were lower than the WHO PV criterion. In these patients, the JAK2V617F allele burden was strikingly similar to that in PV patients fulfilling the 2008 WHO criteria, suggesting that these patients can be classified as PV. Although isotopic measurement of red cell mass (RCM) is required for definitive diagnosis of PV, our data suggest that precise measurement of the JAK2V617F allele burden may improve the diagnosis of PV when RCM has not been determined.

partial, PR (dFLC decrease > 50%) and no response (NR). Patients with baseline and day 100 post-transplant FLC among the CIBMTR cohort were the subjects of this study. Baseline characteristics were assessed. Progression free and overall survival (PFS, OS) were assessed based on the 2012 criteria using the KaplanMeier method. Results: Of 1536 patients, FLC data was available in 104, from 28 North American centers. Median age at transplant was 57 years (Table). Cardiac involvement was reported in 40%; 13% had 4/> organs involved. Majority of patients (71%) had no prior therapy. Day 100 responses were: CR (13), VGPR (65), PR (15) and NR (11). Median follow up was 50 months with 18 deaths (15 from AL). Kaplan-Meier curves for PFS and OS (figure) showed that patients with CR had the best outcomes followed by VGPR. Patients with PR and NR appeared to have equally worse outcomes. Upon combining CR/VGPR versus PR/NR responses, there was a significant difference in OS (p 0.005). Conclusions: In this highly selected AL cohort from a multi-institutional transplant registry, we validate the 2012 criteria. Patients with CR/VGPR have excellent outcomes compared to those with PR/NR. Figure Progression-Free Survival PO-175 The incidence and clinical features of HBV reactivation in multiple myeloma patients treated with novel agents and/or ASCT: a retrospective multicenter study in Japan Y. Tsukune, M. Sasaki, Y. Yahata, H. Tamura, A. Onodera, M. Koike, S. Ito, Y. Ishida, Y. Imai, J. Tanaka, A. Isoda, M. Matsumoto, S. Tanosaki, J. Hua, M. Hagihara, H. Sugimori, T. Odajima, N. Komatsu Department of Hematology, Juntendo University School of medicine; Division of Hematology, Nippon Medical School; Department of Hematology, Juntendo University Shizuoka Hospital; Department of Medical Oncology, Iwate Medical University School of Medicine; Hematology/Oncology, Iwate Medical Department of Hematology, University School of Medicine; Department of Hematology, Tokyo Women’s Medical University; National Hospital Organization Hishigunma Hospital; Department of Hematology, The Fraternity Memorial Hospital; Department of Hematology, Eiju General Hospital; Daito Bunka University. Graduate School of Sports and Health Science, Dept. of Preventive Medicine; Daito Bunka University School of Sports and Health Science, Faculty of Health Science Background: An estimated 2 billion people worldwide have been or are currently infected with hepatitis B virus (HBV), HBV seroprevalence is particularly high in Eastern Asia. HBV reactivation during or after immunosuppressive or cytotoxic therapy has been reported not only in HBsAg positive patients but also in some patients with resolved HBV infection who are negative for HBsAg but seropositive for anti-HBc and/or anti-HBs. Antiviral therapy initiated after hepatitis onset is often insufficient to control HBV reactivation and may lead to death from fulminant hepatitis. In the era of novel agents (bortezomib, thalidomide, and lenalidomide), there have been few reports about HBV reactivation in multiple myeloma (MM) patients and the standard prophylaxis strategy for hepatitis is yet to be established. Patients and Methods: Between January 2006 and June 2014, 99 symptomatic myeloma patients with HBsAg positivity or resolved HBV infection who were treated with novel agents and/or autologous stem cell transplantation (ASCT) were included. Data analyzed included age, gender, M-protein, stage (Durie-Salmon stage, international staging system), laboratory findings, treatment and outcomes using questionnaires. Results: One of 9 HBsAg positive patients developed hepatitis. Among 90 MM patients with resolved HBV infection, HBV reactivation occurred in 15.8% (3 of 19 patients) in the ASCT group and 7.8% (5 of 71 patients) in the non-ASCT group, after 64.5 (7-140) months from the start of chemotherapy; none of them developed hepatitis by means of monitoring HBV DNA levels monthly and administering entecavir. In 5 of these 8 cases, HBV reactivation was noted during bortezomib administration. HBV reactivation has no significant association with age, gender, stage, laboratory findings, and treatment. Conclusion: Since the introduction of rituximab, HBV reactivation has been reported in some malignant lymphoma patients with resolved HBV infection who received rituximab and steroid containing regimen. In the era of novel 15th International Myeloma Workshop, September 23-26, 2015 e183

A 68-year-old female presented with shortness of breath. Chest radiography showed pleural effusion in the right side only. She was suspected of having ovarian cancer because CA125 levels were increased in the pleural effusion, and she consulted our hospital. A chest CT scan showed right pleural nodular lesions. Thoracoscopic pleural resection was performed. Histologic examination of a biopsy specimen showed the diffuse infiltration of small∼medium, mature lymphocytes. These lymphocytes were found to be positive for CD20 and CD79a, but negative for CD3 by immunohistochemistry. These results were interpreted as being consistent with a diagnosis of mucosa-associated lymphoid tissue lymphoma (MALT lymphoma). She commenced chemotherapy with R-CHOP, and the pleural effusion disappeared. MALT lymphoma arising in the pleura is very rare, with only 12 published cases, and most cases have been described in Japan. CA125 levels correlated with the stage, tumor bulk, and presence of effusion. This patient exhibited a high level of CA125 that decreased with therapy.

A 66-year-old male was hospitalized with muscle weakness and gait disturbance. Examination revealed IgM 3,407 mg/dl (IgM, κ-type M protein) and he was diagnosed as having IgM-MGUS neuropathy. He suffered from paralysis of respiratory muscles and required a respirator support. Plasmapheresis and intravenous immunoglobulin were performed and he was weaned from the respirator. Rituximab given as 8 weekly infusions improved gait disturbance. A 71-year-old male was hospitalized with lumbago, numbness of lower extremities and gait disturbance. Examination revealed IgM 1,553 mg/dl (IgM, λ-type M protein) and he was diagnosed with IgM-MGUS neuropathy. Rituximab given as 8 weekly infusions improved gait disturbance. It was concluded that rituximab is a well-tolerated treatment that may be effective in some patients with IgM-MGUS neuropathy.

Introduction: The immunophenotypic analysis of plasma cells using flow cytometry (FCM) is useful for the diagnosis for multiple myeloma (MM) and the detection of MM cells after treatment. CD138 is a well-known surface marker identifying plasma cells, although decreased CD138 expression on plasma cells is frequently observed in MM patients with relapsed or progressive disease. Thus, more stable MM antigens are needed. The SLAM family molecule CD229 has recently been reported to be specifically overexpressed on MM cells including their clonogenic precursors, suggesting that it may be a good marker to identify MM cells. Furthermore, some surface antigens, i.e., costimulatory molecule CD86 (B7-2), its receptor CD28, IL-6 receptor (CD126), insulin-like growth factor-1 (CD221), and coinhibitory molecule B7-H1 (PD-L1, CD274), were reported to be associated with poor prognosis. This study aimed to validate the potential of CD229 as a new MM cell marker and the efficacy of immunophenotypes on MM cells as prognostic markers in a multicenter study. Patients & Methods: Two-hundred thirteen patients comprising 144 newly diagnosed (18 asymptomatic and 126 symptomatic) MM patients, 25 refractory/relapsed MM patients, and 44 monoclonal gammopathy of undetermined significance (MGUS) patients were enrolled. Immunophenotyping of plasma cells in bone marrow was performed with standard 3-color FCM, in which plasma cells were gated by CD38-highly positive cells and 9 parameters, i.e., expression of MM antigens (CD138, CD229) and prognosis-related antigens previously reported (CD28, CD45, CD56, CD86, CD126, CD221, CD274) on MM cells, were analyzed. Expression levels of prognosis-related antigens were compared between patients in high-risk categories, i.e., International Scoring System (ISS) stage II/III, high-risk chromosomal abnormalities [t(4:14), t(14:16), del17p] by FISH, high-risk group stratified by the International Myeloma Working Group (IMWG) [ISS stage II/II and t(4:14) and/or del17p], high serum lactase dehydrogenase (LDH) level, or revised ISS stage III, and other patients. The revised ISS is a powerful new tool to predict outcome in MM patients treated with novel agents based on 3 factors: serum LDH level, ISS stage, and high-risk chromosomal abnormalities [Oliva S, et al. EHA2014, Abstract #S1289]. Differences between continuous variables were evaluated using the Mann-Whitney U-test. Results: 1) CD229 was expressed on almost all MM cells, even on low CD138-expressing MM cells. MM cells from ISS stage II/III patients expressed significantly lower levels of CD138 than those from ISS stage I patients (P = 0.0004). Furthermore, although CD138 expression levels in symptomatic MM patients were lower than those in patients with asymptomatic MM and MGUS and the expression was decreased in refractory/relapsed MM patients, CD229 expression levels on plasma cells were similar in MGUS and MM patients. 2) Newly diagnosed MM patients with high-risk chromosomal abnormalities or in the IMWG high-risk group had higher levels of CD86 and CD126 compared with other patients (P = 0.0056 and 0.0248 in high-risk chromosomal abnormalities, P = 0.0221 and 0.0396 in IMWG high-risk group, respectively). Furthermore, patients in revised ISS stage III had higher levels of CD126 compared with others (P = 0.0646). 3) Although the serum IL-6 level was significantly higher in ISS stage II/III patients, in patients with high LDH levels, and in revised ISS stage III patients compared with other groups, there was no correlation between serum IL-6 levels and expression levels of the IL-6 receptor CD126 on MM cells. Conclusion: CD229 has the potential to become a new marker for the identification of MM cells and target for immunotherapy. High expression levels of CD86 and CD126 were associated with high-risk patients, and CD126 may be associated with survival in patients treated with novel agents. FCM analysis may be useful for predicting prognosis as well as detecting malignant clones. Further studies are in progress to clarify the significance of those molecules on MM cells. Disclosures No relevant conflicts of interest to declare.

CNS recurrence of non-Hodgkin lymphoma is fetal in most cases and tends to occur 5–12 months after the initial diagnosis [1]. Risk factors for CNS involvement include advanced stage, increased IPI, raised LDH, young age, [1 extranodal site, B symptoms, low albumin and retroperitoneal disease. A 19-year-old Japanese man presented to our hospital because of persistent cough on October 2007. A CT scan showed an anterior mediastinal tumor of 8 cm in diameter. LDH value and bone marrow examination were normal. After the total resection, the tumor was diagnosed as primary mediastinal large B-cell lymphoma (PMLBL). Involvement of right infraclavicular lymph nodes detected by Ga-SPECT and FDG-PET/CT determined the stage as IIA. One cycle of MACOP-B regime plus bi-weekly rituximab therapy started on November 2007. After confirming complete response (CR) by CT and Ga-SPECT, 30 Gy of involved field irradiation to the mediastinum and bilateral infraclavicular lymph nodes followed. On June 2008, 3 months after the chemoand radio-therapy, FDG-PET/CT revealed diminishment of abnormal accumulation. On July 2008, only 4 months after the completion of treatment, severe headache and nausea started. A cranial MRI scan showed an enhancing mass in the left frontal lobe with the right midline shift (Fig. 1). Although systemic evaluation revealed no recurrence in the other sites, brain biopsy confirmed infiltration of the lymphoma cells. Southern blot analysis of JH region of immunoglobulin heavy chain gene showed identical rearrangement bands in mediastinal tumor and CNS metastatic mass. The result confirmed that the CNS tumor is truly derived from primary mediastinal mass. The CNS lesion was resistant to two cycles of IVAM therapy containing high-dose methotrexate (MTX) and the following whole brain irradiation of 50 Gy. Then, after obtaining informed consent from the patient, we introduced via Ommaya reservoir three times of 10–25 mg of rituximab together with 3 ml of the autologous serum twice a week. The intracranial rituximab therapy failed to reduce the tumor size at least until day 15, when the patient’s condition rapidly deteriorated and subsequently he died of bacterial pneumonia. PMLBL is a relatively rare subtype of diffuse large B-cell lymphoma (DLBL) and usually occurs in young patients. MACOP-B regimen plus rituximab (R-MACOP-B) combined with local mediastinal radiation ([30 Gy) is recommended as a front-line therapy for the PMLBL patients [2]. This approach has demonstrated a stable progression-free survival in about 70% of the patients 2 years after the diagnosis. The presented patient was also treated with R-MACOP-B combined with involved field radiation and had no CNS risk factors except for young age, but he relapsed only 4 months after the completion of treatment. CNS relapse is reported in more than 10% of NHL arising from the testis, breast and paranasal sinuses, and CNS prophylaxis is recommended to these subtypes of DLBCL [3, 4]. Bishop et al. reported that CNS infiltration occurs in 4% of newly diagnosed PMLBL cases and in 11% of relapsed ones [5]. Although CNS recurrence has been reported in 2–10% of PMLBL patients [6–8], only one report has referred the CNS screening and prophylaxis in PMLBL [3]. Stefoni et al. reported a case of PMLBL M. Sasaki (&) K. Sugimoto A. Masuda Y. Tsukune Y. Yahata N. Komatsu Division of Hematology, Department of Internal Medicine, Juntendo University School of Medicine, 2-1-1 Hongo, Bunkyo-ku, Tokyo 113-8421, Japan e-mail: msasaki@juntendo.ac.jp

Abstract 1972 Classical myeloproliferative neoplasms (MPNs), essential thrombocytosis (ET), polycythemia vera (PV), and primary myelofibrosis (PMF), have a common somatic point mutation, JAK2V617F. Since JAK2V617F heterozygous, homozygous, and wild type cells are randomly and simultaneously detected in the single MPN patient, the quantification of the ratio of JAK2V617F allele to JAK2 total alleles (JAK2 allele burden) would be prerequisite for the assessment of the size of the JAK2V617F-positive clone. Several methods for detecting or quantifying JAK2 allele burden have been developed to date, such as melting peak analysis, and TaqMan Allele Specific PCR (TaqMan AS-PCR). However, these two methods have both pros and cons. The first method can be performed simply to detect the JAK2V617F, but cannot quantify the JAK2 allele burden. The latter is a highly sensitive (0.1 - 1%) method, but requires complicated operations, and the expensive equipment and the reagents. Here, we have established a simple, sensitive, and moderate-in-price method for the quantification of JAK2 allele burden. The method is the combined use of competitive PCR and a sequence-specific fluorescent probe that binds to either the JAK2 wild type gene or JAK2V617F gene. We named the method as alternately binding quenching probe competitive PCR (ABC-PCR). In ABC-PCR, JAK2 wild type and V617F sequences in a target sample are coamplified at the same efficiency using the same primer set; therefore, the allele burden of a target sample does not change before and after the amplification. To measure the allele burden of the target sample, the alternately binding quenching probe (AB-QProbe) is used. AB-QProbe is labeled with a fluorescent dye (BODIPY FL). The fluorescent dye has a property of being notably quenched by an electron transfer to guanine at a particular position. Therefore, the fluorescence intensity is in inverse proportion to the amount of JAK2 wild type (G allele) in a target sample. In other words, the fluorescence intensity reflects the JAK2 allele burden. Thus, the JAK2 allele burden can be simply calculated from the fluorescence intensity of the AB-QProbe at the end-point of PCR. Moreover, ABC-PCR is a highly sensitive method. To determine the lower quantification limit of the allele burden, target samples with known allele burden (50%, 10%, 5%, and 1%) were quantified using ABC-PCR. The experiments were repeated three times, and the mean of the allele burden was calculated. The results of the known allele burden samples (50%, 10%, 5%, and 1%) were 46.9%, 9.5%, 5.7%, and 2.4%, respectively. Furthermore, relative standard deviation (RSD) of each target was also calculated. RSD indicates the deviation of measured values between individual assays. In accurate assays, the value is expected to be less than 0.25. The RSD values for the known allele burden samples (50%, 10%, 5%, and 1%) are 0.05, 0.14, 0.21, and 0.58, respectively; therefore, target allele burden can be accurately quantified until 5% using ABC-PCR. Moreover, the running cost of ABC-PCR is superior to that of TaqMan AS-PCR. The amount of the reagent used in ABC-PCR is reduced to half in comparison with that of TaqMan AS-PCR; therefore, the throughput of ABC-PCR is twice as high as that of TaqMan AS-PCR. Thus, our novel established ABC-PCR assay is the method that possesses simplicity, high sensitivity, and low running cost compared with the existing methods. The assay has a potential to be a golden standard method for the quantification of JAK2 allele burden. Disclosures: No relevant conflicts of interest to declare.