Leukemia, a kind of blood cancer is distinguished by the unregulated growth of malignant white blood cells in the bone marrow, resulting in an excess of aberrant leukocytes. The correct diagnosis and classification of leukemia are critical for efficient treatment and management. Immunophenotyping, a diagnostic approach that detects cells based on their surface antigens, has transformed leukemia diagnosis and treatment. Immunophenotyping allows for the detailed characterization of leukemia cells, enabling the identification of specific sub-types, detection of minimum residual disease, and treatment response monitoring. This method has become a crucial tool in the diagnostic workup of leukemia, supplementing morphological and molecular analysis [1].

A test called immunophenotyping uses the kinds of markers or antigens found on the cell's surface, nucleus, or cytoplasm to detect the cell. The "immuno-" prefix comes from the fact that this method uses antibodies to identify markers or antigens on cells to assist in determining their ancestry. Certain antigens are specific to a single cell type, while others are present on multiple cell types. By contrasting healthy cells with cancerous cells, this method is frequently used to diagnose various forms of leukemia and lymphoma. It is now widely used to identify and categorize acute leukemia, particularly acute myeloid leukemia (AML) [2].

Immunophenotyping is frequently employed in clinical settings to identify hematopoietic cells, but it can also be applied to other types of research. The ability of immunophenotyping to detect minimal residual disease, which enables early intervention and better treatment outcomes, and to identify leukemia subtypes, such as acute myeloid leukemia (AML), acute lymphoblastic leukemia (ALL), chronic lymphocytic leukemia (CLL), and chronic myeloid leukemia (CML), makes it crucial for leukemia diagnosis and management [3]. Regular updates and education for healthcare workers are necessary due to the changing environment of leukemia diagnosis and management, which emphasizes the significance of the current study.

The creation of aberrant leukocytes, which might be a primary or subsequent event, is what defines leukemia. Leukemias are categorized as myeloid or lymphoid depending on the cell of origin and as acute or chronic depending on how quickly the cells proliferate. Acute lymphoblastic leukemia (ALL) and chronic lymphocytic leukemia (CLL), which include the lymphoid lineage, whereas the two most prevalent subtypes, acute myeloid leukemia (AML) and chronic myeloid leukemia (CML), both include the myeloid lineage. Mature white blood cells give rise to other less prevalent variations, including leukemias involving NK cells, mature B-cell and T-cell leukemias, and others. In 2016, the World Health Organization (WHO) revised the designation, drastically changing the conventional classification of acute and myeloid neoplasms [4].

GLOBOCAN, a global observatory for cancer trends, reports that there were 474,519 leukemia cases worldwide, with 67,784 of those instances occurring in North America. The death rate is about 3.2 per 100,000, while the age-standardized incidence rate is about 11 per 100,000. Klinefelter syndrome, Down syndrome, ataxia telangiectasia, Bloom syndrome, and telomeropathies including Fanconi anemia, dyskeratosis congenita, and Shwachman-Diamond syndrome are among the many hereditary risks for leukemia. Additionally, it has been established that the RUNX1 and CEBPA germline genes are mutated.

Major Subtypes of Leukemia

Acute Lymphoblastic Leukemia (All)

ALL, the most widespread leukemia in kids, involves the blastic transformation of B and T cells and accounts for up to 80% of cases in this age group, compared to 20% in adults. It is a highly curable disease in children and leads to treatments derived mainly from pediatric regimens for adolescents (AYA) and young adults with better survival rates.

Acute Myelogenous Leukemia (Aml)

AML is the most widespread type of acute leukemia in adulthood and differs from ALL by morphology and immune marker expression. It is the fastest-growing type of leukemia and response to treatment can change dramatically depending on its molecular subtypes [4].

Chronic Lymphocytic Leukemia (Cll)

CLL often affects individuals between the ages of 60 and 70 y/o and is defined by the clonal maturation of lymphoid cells. It is important to remember that CLL is in general an indolent disease and not the entire CR2 study- group needed treatment because they got symptomatic.

Chronic Myelogenous Leukemia (Cml)

CML usually arises from the reciprocal translocation and fusion of the ABL1 gene on chromosome 9 and the BCR gene on chromosome 22, which results in the Philadelphia (Ph) chromosome, a disrupted tyrosine kinase on chromosome 22. This leads to a monoclonal population of dysfunctional granulocytes, predominantly neutrophils, basophils, and eosinophils.

antigens of a given cell type. Immunophenotyping is usually done via flow cytometry, this is a technique that helps in the rapid determination of several characteristics of a cell. Even though there are major positives with immunophenotyping, it is characterized by many limitations. The complications associated with interpreting results and the likely overlap of shared markers among different leukemia types make the diagnostic process challenging. The strategy is expensive as well, and it could face barriers to accessibility in health systems with limited resources, particularly in certain locations. Notwithstanding this, the merger of immunophenotyping with genomic and molecular studies is creating new avenues for the treatment of leukemia, contributing to the development of customized and more effective therapies. The significant role of immunophenotyping in the study of leukemia's biological variety, the support of treatment choices, and ultimately the enhancement of patient care has become evident with ongoing research progress.

Uses of Immunophenotyping

Immunophenotyping is widely used to distinguish between.

1. Acute myeloid and lymphoid leukemia B and T cell lymphoid neoplasms such as chronic lymphocytic leukemia and lymphoma
2. Reactive and neoplastic expansions of lymphocytes

• Predicting prognosis in lymphoma identification of lymphocyte subsets [3].

Principle of Immunophenotyping

The fundamental principle of immunophenotyping relies on the specific binding of antibodies to antigens on cell surfaces. Antibodies, labeled with fluorochromes, serve as molecular probes, enabling the detection of antigens by flow cytometry or other immunophenotyping techniques. The staining process involves preparing cells, staining with labeled antibodies, and washing away excess antibodies. Standardization is crucial in immunophenotyping, ensuring consistent staining and analysis protocols. Specificity, sensitivity, resolution, and standardization form the foundation of immunophenotyping. Antibody specificity and validation are critical to ensure accurate results [5].

Types of Immunophenotyping Techniques

Flow Cytometry

In flow cytometry, one or more lasers pass over individual cells and particulates floating in a buffered salt-based solution. Visible light scatter by each particulate, one or more fluorescence characteristics are examined. Visible light scatter is quantified in two directions: side (also called side scatter, or SSC), which can indicate the inner complexity or granularity of the cell, and forward (also called forward scatter, or FSC), which may show the relative size of the cell. Fluorescence has no effect on light scatter. Transfection and the generation of fluorescent proteins (such as Green Fluorescent Protein, or GFP), staining with fluorescent dyes (such as Propidium Iodide or DNA), or staining with fluorescently tagged antibodies (such as CD3 FITC) are methods used to prepare samples for fluorescence measurement [6].

Fundamentals of Flow Cytometry

Flow cytometry is the perfect technique for both qualitative and quantitative investigations of the properties of individual cells. It is based on the concepts of light emission, excitation, and scattering. Components of fluorescently tagged cells get excited and release light of different wavelengths when they encounter a laser beam. Fluorescence is utilized to investigate biological characteristics [7].

Flow Cytometry Applications

These are some well-known uses for flow cytometry:

Proteomics in early discovery: One excellent tool for proteomics in early discovery is flow cytometry. Numerous intracellular or cell surface proteins can be evaluated concurrently via assays. Information about protein isoforms can be obtained from the selection of antibodies. Subcellular location or trafficking will be clarified by using an image flow cytometer [8].

Immunophenotyping for cellular composition: Using lineage markers, flow cytometry provides information about a cell population's makeup in immunophenotyping. This aids scientists in identifying unusual cell types and cellular heterogeneity that could be useful for further investigation.

Gene expression analysis in cell signaling pathways: Gene expressions associated with significant cell signaling pathways can be highlighted by flow cytometry. Researchers can make hypotheses about regulatory processes by analyzing stimulated cells for the up- or down-regulation of genes. Genes associated with the phases of the cell cycle are included in this.

MRD using multicomponent flow cytometry (MFC) separates normal cells from leukemia-associated immune phenotypes (LAIP). LAIP is made up of illegitimate markers combined with backbone markers relevant to a cell lineage maturation stage. Numerous institutes have concurrently produced the standard four- to six-color techniques. Consequently, the study protocol determines which marker panels are used. The threshold of detection limit that is regularly attained is 1E-04 [9].

Residual Disease in Acute Myeloid Leukemia (AML)

MRD Detection by PCR: Real-time quantitative PCR (RT-qPCR) is a technique that enables MRD recognition in situations involving chimeric fusion genes produced by balanced chromosomal rearrangements [10].

Immune MRD by Multicolor Flow Cytometry: Identifying leukemia-associated immune phenotypes (LAIP) at diagnosis is the fundamental idea. Despite being modest, the background levels of LAIP in normal and regenerating BM levels hinder the precise detection of abnormalities with sensitivity greater than 1:10,000. The "different from normal" method, which employs a standard fixed antibody panel to identify leukemic cells based on their distinction from normal hematopoietic cells, can be used in the absence of a diagnosis sample [11].

Role of Immunophenotyping

Based on immunophenotyping research, acute leukemias may exhibit unforeseen heterogeneity that cannot be determined by morphological analysis. Numerous investigations have shown that cancerous changes can occasionally produce cellular phenotypes that are both aberrant and transient. The leukemogenic process causes abnormal gene transcription, which is most likely the cause of such unstable phenotypes. This cancerous change in a progenitor cell that might arise along a lymphoid or myeloid routes has therefore been employed to explain lineage heterogeneity. Additionally, it is now clear that the pathophysiology of ALL involves several different molecular pathways and that immunophenotypic subgroups, particularly in B-lineage ALL, reflect a substantial amount of genetic diversity. The ability to distinguish between subgroups that are physiologically and clinically important has been made easier with the use of this knowledge. As a result, immunophenotyping has emerged as a potent tool for characterizing and subclassifying leukemia along with structural and cytochemical investigations by light microscopes. This will aid in understanding the health manifestations of the patient, which could determine the response to treatment [12].

Additionally, immunophenotyping can be helpful when there is little tissue available for a conclusive B-cell lymphoma diagnosis. The expression of CD117 and CD11b in bone marrow has been shown by Rizati et al. to distinguish between recovering benign myeloid proliferation and acute promyelocytic leukemia (APL).

However, contaminated DNA traces usually produce false positive PCR results, which should be taken into account. Clone-specific probes, cautious sample handling and processing, and the use of several controls can all help prevent these mistakes. For the detection of MRD in hairy cell leukemia, flow cytometry is more effective than PCR. It allowed for the quantification of the quantity of tumor cells and was more sensitive and selective. MRD monitoring is becoming more and more crucial for risk-adapted AML patient care. The quantification of MRD levels using multiparameter flow cytometry has demonstrated sensitivity and accuracy with independent prognostic implications.

Prognostic Markers in Leukemia

Prognostic indicators, also known as prognostic factors, are clinical measures used in oncology to assist in determining a patient's probability of a future outcome, such as a recurrence of disease following primary treatment. They are essential to clinical practice because they help differentiate patients into various risk groups, which informs treatment plans and facilitates patient counseling. In order to guarantee treatment group comparability, they can also be used to define strata in clinical trials. Markers can be determined easily as the size of the tumor or the stage of the disease, but they are frequently more complicated, including genetic abnormalities or aberrant protein levels [13].

1. Chronic Lymphocytic Leukemia: In oncology, the idea that clonal mutations may eventually result in the emergence of more hazardous clonal changes is well-established. The clonal evolution of unfavorable aberration over time in CLL is one way it manifests. Due to the extremely low responsiveness of CLL cells to mitogenic stimuli in vitro, only about 50% of patients with CLL exhibit cytogenetic abnormalities using the traditional karyotype banding technique; however, interphase fluorescent in situ hybridization (FISH) can identify aberrations in over 80% of patients. Loss of chromosome 13q14 is the most common anomaly linked to a positive prognosis [14].
2. Acute Myeloid Leukemia: Evaluation of the FLT3 gene mutation status showed 16.7 percent of the patients had FLT3-internal tandem duplication (ITD) mutation, while others had wild-type FLT3. The qRT-PCR technique was utilized to evaluate the manifestation levels of the four genes that were selected. ASXL1 expression levels in AML patients did not differ substantially from those in normal subjects; transcript levels in 28 patients (52%) showed an elevated level with an RQ range of 1.62 to 16, while in 13 patients (24%), there was a decrease in RQ range of 0.065 to 0.9.

• Chronic Myeloid Leukemia: The main function of MFG-E8 (Milk Fat Globule–Epidermal Growth Factor-8) is to synthesize the membrane glycoprotein cadherin, which facilitates the killing of apoptotic cells. MEG-E8 stimulates invasion, immunological suppression, and the epithelial-to-mesenchymal transition in melanoma cells, while its high expression is associated with the advancement of oral, breast, prostate, and colon malignancies. Since rs4945 was implicated in the development of numerous malignancies, it was also anticipated that it would be linked to the failure of IM treatment in CML. The alpha subunit of C8 is encoded by complement component 8-alpha (C8A). The membrane attack complex (MAC) is formed in part by C8A. In patients with breast cancer receiving trastuzumab, raised RNA levels of the C8A gene led to a good prognosis for hepatocellular carcinoma.

1. Acute Lymphocytic Leukemia: In the last few decades, the prognosis for ALL has significantly improved due to improvements in supportive care, optimization of the current chemotherapeutic medications, and therapy adaptation to the degree of relapse risk. Children's outcomes have significantly improved over recent decades, with long-term survival rates approaching 80% and total remission nearing 95%. Adults with ALL continue to receive inadequate therapy, unlike children with ALL. With a 5-year overall survival of only 30–40% for patients under 60, fewer than 15% for those over 60, and less than 5% for those over 70, the long-term survival rate for adults has not altered much over the last 20 years [15].

Immunophenotyping in Monitoring Leukemia Treatment

The Application of Cell Surface Markers:

The identification of cell surface antigens by monoclonal antibodies (MoAbs) has provided crucial information about leukocyte differentiation and the cellular cause of leukemia. It is now feasible to precisely identify stages of human lymphocyte and granulocyte differentiation using those highly specific MoAbs because leukemic cells display several leukocyte differentiation antigens that reflect commitments to the myeloid or lymphoid lineage as well as levels of maturation. Groups of MoAbs that identify the same antigen have been recognized as cluster designations or clusters of distinct ion antigens (CDs) as an outcome of the International Workshops on Human Leukocyte Differentiation Antigens. Nowadays, cell lineage and maturation stage are defined by these CDs [16]. Normal marrow B cells were separated into four distinct developmental stages by cell membrane by Loken et al.

Leukemia Associated Antigens

Leukemia-associated antigens (LAAs) are being increasingly identified by methods such as cytotoxic T-lymphocyte (CTL) cloning, serological analysis of recombinant cDNA expression libraries (SEREX), and mass spectrometry (MS). The role of tumor antigens in the biological processes that are disrupted in acute myeloid leukemia (AML) has also been better understood with the aid of large-scale screening methods like microarray, single nucleotide polymorphisms (SNPs), serial analysis of gene expression (SAGE), and 2-dimensional gel electrophoresis (2-DE). These antigens serve as both immunotherapy targets and indicators of disease status, stage, response to treatment, and survival. Biomarkers are necessary to help physicians and scientists enhance clinical outcomes and treatment design, as well as to identify which patients are most likely to benefit from particular treatments, whether they are traditional or innovative [17].

Nonetheless, certain genes are overexpressed in AML and linked to particular AML subtypes, such as PRAME. Patient CTLs were the first to identify PRAME as an antigen in human melanoma [18].

Acute Leukemia: Acute leukemias are typically managed as inpatient cases requiring intensive support, frequent monitoring, and management of complications such as infections and electrolyte imbalances. Key considerations at diagnosis include identifying acute promyelocytic leukemia (APL), as it has a distinct treatment protocol compared to other acute myeloid leukemia (AML) subtypes.

Acute Promyelocytic Leukemia (Apl):

A hemorrhaging diathesis associated with reduced fibrinogen and high coagulation parameters (PT, aPTT) is indicative of APL (Indunil et al. 2024). Immediate Treatment: Initiate all-trans-retinoic acid (ATRA) upon suspicion of APL to promote the differentiation of pro myeloblasts into mature granulocytes, thereby preventing complications such as differentiation syndrome. Differentiation Syndrome: Manifests with fever, respiratory distress, and edema. Treated with dexamethasone at 10 mg every 12 hours until symptom improvement [4].

Monitoring: Frequent electrolyte checks and ECGs to monitor QTc prolongation during ATO therapy.

Acute Myeloid Leukemia (AML):

Standard Therapy: The '7+3' regimen, which consists of a 3-day treatment of an anthracycline and a 7-day continuous infusion of cytarabine (daunorubicin or idarubicin) [4].

Risk Stratification: Based on cytogenetic and molecular markers, guiding prognosis and treatment decisions [4].

Minimal Residual Disease (MRD): Monitoring MRD levels post-induction therapy to guide further treatment, including bone marrow transplantation (BMT) [4].

4.2 Chronic Leukemia

Chronic Myeloid Leukemia (Cml):

Targeted Therapy: Tyrosine kinase inhibitors (TKIs) targeting the Philadelphia (Ph) chromosome.

Resistance Management: Testing for mutations (e.g., T315I) and using alternative agents like ponatinib, asciminib, or omacetaxine, with SCT as a last resort for multiple TKI resistance [4].

Chronic Lymphocytic Leukemia (Cll):

Indolent Course: Treatment is initiated based on specific criteria such as rapid lymphocyte doubling time, worsening cytopenias, splenomegaly causing discomfort, and significant B symptoms.

Prognostic Factors: IGVH mutation status, del17p, TP53 mutation, and t (11:14) to rule out mantle cell lymphoma [4].

• Good Prognosis (IGVH mutation): Chemotherapy regimens such as FCR (fludarabine, cyclophosphamide, rituximab) or BR (bendamustine, rituximab) can achieve prolonged disease-free survival.
• High-Risk (del17p/TP53 mutation): Targeted therapies like venetoclax (BCL-2 inhibitor) or BTK inhibitors (ibrutinib, acalabrutinib), often in combination with rituximab or obinutuzumab.

Richter Transformation: Rapid progression of CLL/SLL to aggressive diffuse large B-cell lymphoma, requiring urgent biopsy and tailored treatment.

These treatment protocols emphasize the importance of personalized care based on the specific leukemia subtype and individual patient factors, ensuring optimal outcomes and management of the disease [4].

Differential Diagnosis: Because leukemia symptoms are non-specific, the differential diagnosis is wide. Other illnesses that can cause similar anomalies in blood cell counts must be ruled out. Examine the possible differential diagnosis listed below:

Deficits in vitamins and micronutrients, such as deficits in copper, folate, and B12.

Viral infections, including Epstein-Barr virus, CMV, and HIV Drug effects include ganciclovir, mycophenolate mofetil, valproic acid, and chemotherapy drugs.

Systemic lupus erythematosus (SLE) is an example of an autoimmune condition.

Prognosis: The prognosis is significantly influenced by the patient's age, concomitant disorders, cytogenetic and molecular findings, and leukemia subtypes. In general, the 5-year survival rate for leukemia has risen over time, from 33% in 1975 to 59% in 2005 [4].

4.3 Complications:

Complications: Acute promyelocytic leukemia is frequently linked to disseminated intravascular coagulation (DIC), an acute condition that can occur in other leukemia subtypes as well. DIC is caused by a malfunction in the blood clotting proteins, which can result in both thrombosis and bleeding. To avoid serious cardiac toxicity, aggressive fibrinogen replacement with cryoprecipitate and routine laboratory monitoring are advised.

A dangerous consequence known as disseminated intravascular coagulation (DIC) occurs when the blood clotting proteins malfunction, resulting in thrombosis and bleeding. Although it can happen in other leukemia subtypes, acute promyelocytic leukemia is frequently linked to it.

Management: Active fibrinogen replacement with cryoprecipitate and regular laboratory monitoring are essential for patient survival.

Infection: Patients with immunosuppression from chemotherapy, stem cell transplantation, or leukemia itself are at a higher risk of developing serious infections. When an immunocompromised patient has a fever and neutropenia, broad-spectrum antibiotics should be started right away [4].

1. Complexity of Results: To distinguish between healthy and diseased cell populations, immunophenotyping data interpretation can be challenging and calls for skilled professionals.
2. Overlap of Markers: Certain leukemias share immunophenotypic markers, which could lead to misidentification or make it harder to distinguish between distinct types.
3. Lack of Functional Details: While immunophenotyping aids in the identification of different cell types, it is completely useless in explaining the functions of the cells, which is essential for determining the progression of diseases.
4. Availability: Not all areas may have access to advanced immunophenotyping technologies, which could be concerning and have an impact on the legitimacy of different treatment and diagnostic approaches. It is important to note that every patient has an equal right to new treatment options.
5. Cost and Resources: In general, sophisticated immunophenotyping methods are costly; this may be a barrier to their widespread use, particularly in environments with limited resources [19]. Because of these limitations, it is clear that supplementary testing approaches, such as genomic and molecular testing, are desperately needed to improve the accuracy and efficacy of leukemia care.

Future Direction

Flow cytometry has been used for decades to determine the presence and individual proportions of particular leukocyte groups, a process known as immunophenotyping. Immunophenotyping is used in clinical settings to detect autoimmune disorders, autoimmune deficits, and hematological cancers. Consequently, the most effective cutting-edge technique for identifying relevant extracellular and intracellular proteins at the single-cell level is flow cytometry. Up until recently, quick protein level detection and simple downstream analysis of the produced data were made possible by the binding of fluorophore-labeled antibodies to their targets. However, the restricted number of detectors used in conventional flow cytometry leads to compensatory errors and fluorescent overlap of the various fluorescent dyes. Spectral flow cytometry was established as a result of recent developments [20].

The full emission spectra of individual fluorophores across all lasers may be measured thanks to these innovative multi-detector techniques. Because each fluorophore has a unique spectral fingerprint created by full spectrum technology, dyes with nearly identical emission peaks can be mathematically separated, something that is not possible with traditional flow cytometry. The ability to extract the autofluorescence of cells, which leads to higher population resolution of samples, is an additional benefit over the traditional approach. Additionally, by lowering the signal spreading error, spectral flow cytometry shows improved population separation ability. As a result, full-spectrum technology makes it possible to create huge, flexible staining panels that can add more, highly overlapping fluorophores to increase the application range. A 40-color assay was recently established in the rapidly developing field of spectral flow cytometry [20].

Immunophenotyping provides information on the type of leukemic cell and its maturation status which are essential in recognizing a particular kind of leukemia, its progression, and reaction to treatments.

1. Arber DA, Orazi A, Hasserjian R, Thiele J, Borowitz MJ, Le Beau MM et al. The 2016 revision to the World Health Organization classification of myeloid neoplasms and acute leukemia. Blood. J American society hematology. 2016; 127: 2391-405.
2. Chen DC, Potok OA, Rifkin D, Estrella MM. Advantages limitations and clinical considerations in using cystatin c to stimate GFR. Kidney 360. 2022; 3: 1807-1814.
3. Yan B, Gong B, Wang X, Zheng Y, Sun L, Wu X. et al. Embryonic lethal phenotyping to identify candidate genes related with birth defects. Int J Mol Sci. 2024; 25: 8788.
4. Indunil M. Hewage H, Nye DH, Elissa J, Schwartz S. How does vaccine-induced immunity compare to infection-acquired immunity in the dynamics of covid-19. Pathogens. 2015; 14: 179.
5. Araki D, Wood BL, Othus M. Allogeneic hematopoietic cell transplantation for acute myeloid leukemia: time to move toward a minimal residual disease-based definition of complete remission? J Clin Oncol. 2016; 34: 329-336.
6. Bader P, Kreyenberg H, Stackelberg A. Monitoring of minimal residual disease after allogeneic stem-cell transplantation in relapsed childhood acute lymphoblastic leukemia allows for the identification of impending relapse: results of the ALL-BFM-SCT 2003 trial. J Clin Oncol. 2015; 33: 1275-1284.
7. Bader P, Kreyenberg H, Henze G. Prognostic value of minimal residual disease quantification before allogeneic stem-cell transplantation in relapsed childhood acute lymphoblastic leukemia: the ALL-REZ BFM Study Group. J Clin Oncol. 2009; 27: 377-384.
8. Darzynkiewicz Z, Halicka HD, Zhao H. Analysis of cellular DNA content by flow and laser scanning cytometry. Adv Exp Med Biol. 2010; 676: 137-147.
9. Pedreira C, Costa E, Lecrevisse Q. Overview of clinical flow cytometry data analysis: recent advances and future challenges. Trends Biotechnol. 2013; 31: 415-425.
10. Grimwade D, Freeman SD. Defining minimal residual disease in acute myeloid leukemia: which platforms are ready for prime time? Blood. 2014; 124: 3345-3355.
11. Loken MR, Alonzo TA, Pardo L, Gerbing RB, Raimondi SC, Hirsch BA, et al. Residual disease detected by multidimensional flow cytometry signifies high relapse risk in patients with de novo acute myeloid leukemia: a report from Children's Oncology Group. J American Society Hematology. 2012; 120: 1581-1588.
12. Ganggaiswari A, Kresno SB, Krisnuhoni E. VEGF expression and desmoplastic reaction as potential progressive factors in young patients with colorectal cancer. Acta Med Indones. 2010; 42: 6-11.
13. Riley JL, June CH, Blazar BR. Human T regulatory cell therapy: take a billion or so and call me in the morning. Immunity. 2009; 30: 656-665.
14. Mousa A, Al-Taiar A, Anstey NM, Badaut C, Barber BE, Bassat Q, et al. The impact of delayed treatment of uncomplicated P. falciparum malaria on progression to severe malaria: A systematic review and a pooled multicentre individual-patient meta-analysis. PLoS Med. 2020; 17: e1003359.
15. Mirela Cristina L, Matei D, Ignat B, Popescu CD. Mirror therapy enhances upper extremity motor recovery in stroke patients. Acta Neurol Belg. 2015; 115: 597-603.
16. Kresno SB, Haryanto SH, Kosasih AS, Muthalib A, Atmakusumah D. Immuno phenotyping in leukemia and its diagnostic significance. Med J Indonesia. 2004; 13: 195-202.
17. Guinn DA, Abel DE, Tomlinson MW. Early goal directed therapy for sepsis during pregnancy. Obstetrics and gynecology clinics of North America. 2007; 34: 459-479.
18. Ikeda H, Ohta N, Furukawa K, Miyazaki H, Wang L, Furukawa K. Mutated mitogen-activated protein kinase: a tumor rejection antigen of mouse sarcoma. Proceedings of the National Academy of Sciences. 1997; 94: 6375-6379.
19. Wang Y, Jodoin PM, Porikli F, Konrad J, Benezeth Y, Ishwar P. CDnet 2014: An expanded change detection benchmark dataset. In proceedings of the IEEE conference on computer vision and pattern recognition workshops 2014; 387-394.
20. Preglej T, Brinkmann M, Steiner G, Aletaha D, Göschl L, Bonelli M. Advanced immunophenotyping: A powerful tool for immune profiling drug screening and a personalized treatment approach. Front Immunol. 2023; 14: 1096096.