Molecular laboratory investigations for myeloproliferative neoplasm (MPN) can ideally be divided into two distincts groups, those for the detection of the BCR-ABL rearrangement (suspect of chronic myeloid leukemia) and those for the variants determination of the driver genes of the negative Philadelphia forms (MPN Ph neg).

The BCR-ABL detection is based on RT-Polymerase Chain Reaction techniques and more recently on droplet digital PCR (ddPCR). For this type of analysis, combined with chromosome banding analysis (CBA) and Fluorescent in situ hybridization (FISH), it is essential to quantify BCR-ABL mutated copies by standard curve method.

The investigation on driver genes for MPN Ph neg forms includes activity for erythroid forms such as Polycythemia Vera (test JAK2V617F and JAK2 exon 12), for non-erythroid forms such as essential thrombocythemia and myelofibrosis (test JAK2V617F, CALR exon 9, MPL exon 10), for “atypical” ones such as mastocytosis (cKIT D816V test) and for hypereosinophilic syndrome (FIP1L1-PDGFRalpha test). It's crucial to assign prognosis value through calculating allelic burden of JAK2 V617F variant and determining CALR esone 9 variants (type1/1like, type2/2like and atypical ones).

A fundamental innovation for investigating triple negative cases for JAK2, CALR, MPL and for providing prognostic score is the use of Next Generation Sequencing panels containing high molecular risk genes as ASXL1, EZH2, TET2, IDH1/IDH2, SRSF2. This technique allows to detect additional or subclonal mutations which are usually acquired in varying sized sub-clones of hematopoietic progenitors. These additional variants have a prognostic significance and should be indagated to exclude false negative cases.

骨髓增生性肿瘤 (MPN) 的分子实验室研究可以理想地分为两个不同的组，用于检测 BCR-ABL 重排（疑似慢性髓细胞白血病）的组和用于确定费城阴性形式的驱动基因变异的组（MPN Ph 负值）。

BCR-ABL 检测基于 RT-聚合酶链式反应技术，最近基于液滴数字 PCR (ddPCR)。对于此类分析，结合染色体显带分析 (CBA) 和荧光原位杂交 (FISH)，必须通过标准曲线法量化 BCR-ABL 突变拷贝。

对 MPN Ph 阴性形式的驱动基因的研究包括对红细胞形式如真性红细胞增多症（测试 JAK2V617F 和 JAK2 外显子 12）的活性，对非红细胞形式如原发性血小板增多症和骨髓纤维化（测试 JAK2V617F、CALR 外显子 9、MPL 外显子 10）的活性），用于“非典型”的，如肥大细胞增多症（cKIT D816V 测试）和嗜酸性粒细胞增多症（FIP1L1-PDGFRalpha 测试）。通过计算 JAK2 V617F 变体的等位基因负荷和确定 CALR esone 9 变体（type1/1like、type2/2like 和非典型变体）来分配预后价值至关重要。

研究 JAK2、CALR、MPL 三阴性病例和提供预后评分的一项基本创新是使用下一代测序面板，其中包含 ASXL1、EZH2、TET2、IDH1/IDH2、SRSF2 等高分子风险基因。该技术允许检测通常在不同大小的造血祖细胞亚克隆中获得的额外或亚克隆突变。这些额外的变体具有预后意义，应该排除假阴性病例。