Harmonization of laboratory hematology: a long and winding journey

The harmonization of diagnostics in hematology is an issue whose complexity parallels that of laboratory hematology. This area of diagnostic testing conventionally entails a combination of first-line tests, mainly represented by the complete blood cell count (CBC), and second-line analyses which can be based on different techniques such as cytochemistry, cytogenetics, cytofluorimetry, up to genomics and transcriptomics. Throughout the broad area of laboratory hematology, and thus starting from the “basic” CBC, widespread standardization and harmonization almost remain unmet targets [1], [2], [3], [4] (Table 1).

Due to the many technological advancements occurred over the past few years, laboratory hematology has undergone a considerable transformation, which has also entailed the possibility to perform morphological analysis of peripheral blood smear through digital optical microscopy and generate many new blood cell population parameters, also including the so-called cell population data [1], [2]. More specifically, innovative technologies and analytical principles for morphological analysis, along with continuous evolution of hardware and software, have allowed to manufacture a new generation of hematological analyzers (i.e. “hemocytometers”), which are now capable to produce both quantitative (i.e. cell counts and cell indices) and qualitative (i.e. cytograms and morphological alarms) parameters [1], [2]. These important features have led the way to developing diagnostic algorithms, often completely automated, which will better suit the modern organization and volume of clinical laboratories. Nevertheless, these remarkable progresses have also generated new threats, such as the need of increasingly qualified knowledge for better identifying the many potential clinical applications, along with the compelling necessity to manage the potential inappropriateness of (innovative) test requests.

Depending on the different technology, the new hemocytometers are now capable to generate a vast array of innovative parameters (e.g. nucleated red blood cell count, immature granulocytes [IG] count, immature platelet [IPF] and reticulocyte fractions, mean reticulocyte hemoglobin content [RET-He]) along with more traditional tests, whose clinical significance needs to be clearly acknowledged. On the other hand, the increasing technological heterogeneity characterizing the different hematological analyzers can be seen as a serious threat for harmonization, thus complicating further the potential clinical usefulness of hematological test results [1], [2], [4].

Literature data shows that the some basic parameters of the CBC count have now achieved a reasonable inter-instrumental agreement, whilst important limitations persist for monocyte and basophil counts, as well as for the assessment of some instrument-dependent erythrocyte and platelet indices [1], [2], [4]. The heterogeneity of the analytical performances further emphasizes the need to invest larger efforts in standardization and harmonization of these parameters, especially when used in the diagnostic approach of patients with thrombocytopenia. Better standardization and harmonization should also be achieved for automated reticulocyte count, for measurement of reticulocyte indices and for assessment of more specific parameters such as IG or IPF [2], [4].

Scarce information is currently available on standardization and/or harmonization of all the over-20 basic parameters included in the CBC, i.e. hemoglobin, hematocrit (Ht), erythrocyte count, leukocyte and platelet counts; erythrocyte indices (e.g. mean corpuscular volume, mean corpuscular hemoglobin [MCH] and mean corpuscular hemoglobin concentration [MCHC]); and white blood cell differential with five leukocytes populations (i.e. neutrophils, lymphocytes, eosinophils, basophils and monocytes). This is mainly due to the fact that the assessment of these parameters is mostly technique and instrument dependent. The essential drawbacks hurdling harmonization of some “conventional” calculated (i.e. MCHC, MCH and Ht) and measured (e.g. mean platelet volume and red blood cell distribution width) parameters are well known, and it is hence no surprising that the challenges for harmonizing more complex and innovative tests such as IPF, RET-He and IG will be exponentially bigger [1], [2], [4]. A guideline for hemocytometers evaluation has recently been published by the International Council for Standardization of Hematology (ICSH) [5], in the attempt to overcome some of these hurdles.

The qualitative information generated by the hemocytometers, such as morphological flags and other alarms, is plagued by the highest level of heterogeneity because they are highly technique and instrument dependent [4], [16]. The modest efforts made so far for harmonizing the flags associated with morphological lymphocyte abnormalities are especially concerning. For example, the description used for an identical morphological flag (i.e. “atypical lymphocytes”) has often distinctive significance in different analyzers. The generation of this alarm may hence mirror the presence of neoplastic lymphocytes in some hemocytometers, and it may be associated with the presence of reactive lymphocytes (e.g. as in the case of some viral infection) with other instrumentation. Notably, no quality control programs have become available so far for verifying the analytical performance over time of qualitative parameters, such as morphological flags.

The test results produced by some recent hematological parameters are unfortunately plagued by poor comparability, and this is especially alarming because they may convey valuable clinical information, are almost inexpensive and are available 24 h a day. It is hence advisable that additional harmonization efforts should be planned to verify their clinical applicability, albeit the different technologies and the high commercial competition will not facilitate this process.

In 2017, many new hemocytometers and software upgrades have become commercially available, thus reflecting the high level of commercial competition in this area of diagnostic testing. Interestingly, although newer analyzers are in the pipeline, with the capability of generating an even larger number of innovative and increasingly useful parameters, there is little evidence that the manufacturers are actively involved in harmonization campaigns.

Albeit morphological analysis of peripheral blood smear retains an essential role for diagnosing hematological disorders, the competency of laboratory medicine professionals is often inadequate. This can be seen as either a cause or a consequence of insufficient agreement between diagnostic pathways and the so-called reflex testing criteria used for triggering microscopic revision of blood smears after automated analysis has been completed [17], [18].

Regarding morphological characteristics of blood cells, many resources have been invested by the scientific community for harmonizing nomenclature and for standardizing optical analysis, such as the suggested classification of acute leukemia published by the French-American-British working group [19]. The working groups and task forces currently active in this area include the ICSH [6] the European LeukemiaNet network (with the project “An atlas of cells with consensual nomenclature by the WP10 Morphology Faculty”) [6] and the working group on Morphology of Myelodysplastic Syndrome [7], [8], [9], [10].

The recent development of innovative analytical systems for performing digitalized morphological analysis of peripheral blood smear discloses unique but challenging scenarios, such as the need to verify and monitor their analytic performance, as well as the definition of harmonization pathways for digital morphological assessment [3].

These important challenges reinforce the need of promoting the organization of qualification courses for specialists in laboratory medicine (in particular for morphological assessment of peripheral blood smears), as well as for fostering the integration of these skills with innovative techniques and/or methods, especially with immunological and molecular testing.

The process of test results communication, whose accuracy is essential for managed care of both hematological and non-hematological disorders, may also benefit from a better level of harmonization [12], [13], [14], [15]. The general requirements of laboratory reports are clearly specified in the International Organization for Standardization 15189 standard, whereas some of those related to hematologic tests reporting can be found in the scientific literature, especially for quantitative parameters, measurement units and management of critical values [12], [13] (Table 1). However, only little information is now available for the minimum reportable information (i.e. number and type of quantitative parameters), as well as for qualitative comments [14], [15]. Although the CBC is primarily based on quantitative parameters, the results of some of these tests may still benefit from being accompanied by comments regarding both quantitative and qualitative (i.e. morphological) abnormalities, which are often present in association with hematological and non-hematological disorders.

The harmonization of comments in laboratory reports is another essential area, because these expert advices will provide an essential guidance to the clinical decision making. Two specific regional projects have been very recently started in this area, the former from the French-Speaking Cellular Hematology Group [14] and the latter from the working group on Diagnostic Hematology of the Italian Society of Clinical Chemistry and Clinical Molecular Biology [15].

In conclusion, the remarkable technical advancements recently occurred in laboratory hematology [20], along with the appealing perspective of major integration with cytofluorimetry, digital analysis and molecular biology, will need to be supported by continuous education and training of laboratory professionals. Scientific societies and organizations should also plan larger efforts for harmonizing the entire diagnostic hematology process [21]. In this perspective, the development of integrated laboratory reports, containing information gathered from basic hematological tests (e.g. CBC) and more complex investigations (i.e. molecular biology, immunohematology), shall be seen as an ambitious but profitable target [22].