Super Resolution Localization Microscopy Enables Quantitative Diagnosis of Multiple Myeloma
Authors: Tong Xiqin , Kuang Weibing , Liang Yuxing , Zhou Fuling , Huang Zhen li 
Published: November 2021

Abstract
Multiple myeloma (MM) is a bone marrow-based malignancy that has a range of consequences resulting from either its direct effects on the bone marrow microenvironment- causing anaemia, more extensive myelosuppression and bone lysis - or its indirect effects on the kidney and other organ systems. Over the past several years, the introduction of autologous stem cell transplantation, immunomodulatory drugs, proteasome inhibitors, histone deacetylase inhibitors, and monoclonal antibodies has substantially improved survival outcomes. However, MM remains biologically heterogeneous with significant variability among patients in terms of clinical features, response to therapy and overall survival (OS). Several prognostic variables can help predict this variability in outcomes ranging from clinical based systems such as the International Staging System (ISS) to more advanced molecular characterizations of the myeloma PCs by cytogenetics and gene expression profiling. However with the advancement in technology utilized for laboratory testing and the emergence of new treatments for MM, there has been an evolution in the significance of these previously defined prognostic markers with time. Assessment of disease activity and depth of response continues to be a moving target in MM. Super-resolution fluorescence imaging is a major breakthrough in the field of optical imaging in this century. Super-resolution fluorescence imaging has been applied to a variety of biological imaging applications, including membrane, cytoskeletal and cytosolic proteins in fixed and living cells. Molecular motions can be quantified.
To establish the detection limit and sensitivity threshold of dSTORM and FC, we used serial dilutions of anti-CD38 antibody to detect expression of CD38 on 8226 cells by Super Resolution localization Microscopy and flow cytometry (FC). Design six different antibody concentrations (300ng/ml, 30ng/ml, 10ng/ml, 3ng/ml, 1ng/ml, 0.3ng/ml) to label MM cells with immunofluorescence, and then detect them by flow cytometry. Similarly, design eight different antibody concentrations (1μg/ml, 300ng/ml, 30ng/ml, 10ng/ml, 3ng/ml, 1ng/ml, 0.3ng/ml, 0.1ng/ml), and perform the same treatment, perform immunofluorescence labeling on MM cells, and then perform super-resolution imaging, and calculate the density of CD38 protein on the surface of MM cells of each concentration. Figure 1A detects the ratio of the number of cells to the total number of cells, it can be seen from Figure 1A that when the antibody concentration is not less than 30ng/ml, almost all MM thinners can detect the signal, but when the antibody concentration is less than at 30ng/ml, only a few or no cells can detect the signal. This shows that when the antibody concentration is lower than 30ng/ml, the sensitivity of flow cytometry is no longer sufficient to detect CD38 protein on the surface of MM cells. From Figure 1B, it can be seen that even when the CD38 antibody concentration is 0.3ng/ml, super-resolution imaging can still accurately identify each CD38 antigen molecule on the surface of MM cells, and the statistical CD38 protein density is 8.5864±1.4180/μm2. This shows that the sensitivity of super-resolution is much higher than that of streaming.

Disclosures
No relevant conflicts of interest to declare.