Combining Technologies To Separate Blood Cells Efficiently

Optical forces can be applied from the outside of the microfluidic device, allowing development of highly modular, multi-purpose systems for cell sorting.

Human blood is an essential component of human life which is universally recognized as the most precious element that sustains life. Till today, there is no substitute of human blood. Though under trial, placental, cadaveric, and artificial blood are not fulfilling the desired parameters and cannot be established as an alternate source of whole human blood. Transfusion medicine has undergone diverse and fascinating advancements since its initiation in the early 20th century. One of these was the discovery that blood can be divided into individual components and delivered separately. Availability of safe blood and blood products is a critical aspect of improving healthcare. Adherence to proper indications for blood component therapy is essential because of its potential adverse effects and costs of transfusion. Over the years, the significance of blood components in treating certain diseases or conditions has been recognized. In the fields of biomedicine and biological research, efficient and high-throughput cell sorting and manipulation are crucial. Cell sorting and manipulation methods development and miniaturization are key to point-of-care diagnostics and therapeutics research.

In biomedical research and clinical diagnostics, along with filtration, centrifugation, and sedimentation techniques, fluorescent activated cell sorting (FACS) and magnetic activated cell sorting (MACS) have become standard methodologies for accurate and continuous sorting of physically similar, heterogeneous mixtures of cells and particles. FACS and MACS technologies have reached maturity, so their improvement to achieve lower cost, higher portability, smaller sample sizes, and greater purity has become a difficult task. These factors have led many researchers to study alternative methods of cell separation such as acoustic separation and buoyancy-activated cell separation (BACS). In the field of clinical diagnostics and therapeutics, novel, miniature separation technologies are allowing to achieve extremely high precision of cancer diagnostics. In the case of blood cells, diagnosis of many diseases requires extraction and analysis of specific blood cell populations, such as erythrocytes, leukocytes, platelets, and pure plasma.

Cell sorting is also being extensively used in regenerative medicine and tissue engineering applications. Another emerging field of study where cell separation techniques are being employed is personalized medicine, where rapid and accurate cell separation and enrichment is of paramount importance. Finally, fundamental biological studies focusing on understanding of individual characteristics of various populations of cells make heavy use of novel cell separation platforms. Furthermore, the modern day transfusion medicine has the ability to provide transfusion products specific to the patient’s need which greatly improves treatment outcomes and maximizes the donor resources. Separation of whole blood into its constituents allows optimal survival of each component. This ability to transform whole blood into various components relies heavily on good centrifuges and component separators.

Indian scenario
With more than 1200 road accidents occurring every day in India, 60 million surgeries, 240 million major operations, 331 million cancer-related procedures like chemotherapy, and 10 million pregnancy complications all require a serious call for transfusion of blood and blood products. Therefore, a well-organized, well-structured, and effective blood transfusion service is vital for the healthcare delivery system. Blood and blood products are a precious commodity which gives life to another person. Though immense discoveries and inventions have been made in science and technology, yet one cannot make blood; hence, human blood has no substitute.

The availability of safe blood and blood products is essential for diverse modern healthcare services including some surgeries, treatments for cancer, chronic medical conditions, trauma care, organ transplantation, and childbirths that ultimately improve the lives of millions of patients who are in need of transfusion annually. Still, the country has not yet developed a well-defined and stringent regulatory framework for blood products regulation. Frailty may arise from the inability of governments to enforce laws, regulations, and policies and personnel who may not be aware or cannot follow quality assurance and/or good manufacturing practices (GMP).

At present, India has 2903 blood banks spread all across the country, of which 1043 are public and 1860 private, including those run by charitable trusts. Maharashtra has 328 (the most), followed by Uttar Pradesh (294) and Tamil Nadu (291). On the other hand, 74 districts across 17 states do not have a single blood bank. Assam has 12 such districts, followed by Arunachal Pradesh and Telangana, each with 10. To overcome these shortcomings, the government has planned to set up blood banks in 68 districts of the country to provide services in the rural hinterland. With the constant effort of government and other organizations to increase blood donations there is a need for blood cell separators and other equipments in every blood bank to overcome the requirement of blood components which increases with incidences like a dengue outbreak. All this will lead to increase in the market for blood cell separators and centrifuges.

Several installations of apheresis machines are taking place in the country. For instance, the Andhra Pradesh Health Minister, Kamineni Srinivas in keeping with the promise to improve facilities at the Rajiv Gandhi Institute of Medical Sciences (RIMS) recently inaugurated a blood component separator facility at the RIMS blood bank. Similarly, the Ooty government hospital has installed a blood component separator. The Indira Gandhi Institute of Medical Sciences (IGIMS), Patna, Bihar is planning an apheresis machine and the MGM Medical College and Hospital, Jamshedpur, Jharkhand also plans to acquire a blood component separator for its blood bank.

Global scenario
The global apheresis market is projected to gain humongous revenue of USD 4.33 billion by 2025, from initial revenue valued at USD 1.85 billion in 2017, growing at a CAGR of 10.2 percent, according to Transparency Market Research. Widespread prevalence of chronic diseases all over the world has caused a high demand to exist regarding use of various blood components such as plasma, albumin, immunoglobin, and other coagulation factors. Such a high demand is one of the primary reasons for the global apheresis market to exhibit excellent growth. Extraction of components is mainly done using apheresis. Also, strict regulation for maintaining safety in various healthcare facilities has made specialists employ apheresis methods, thereby propelling the market to pick up substantial pace. With rapid advancements occurring in the field of therapeutic services, many healthcare organizations have made notable progress in the field of blood component extraction, consequently proving to be beneficial to the market.

However, this market is being restrained mainly due to shortage of availability of relevant equipment and other accessories required to perform necessary procedures in remote regions. High cost of the equipment might be passed onto the patients and their kin in the form of exorbitant medical fees. This could discourage people mainly from underdeveloped and developing regions from opting for apheresis, primarily due to presence of less disposal income. Nevertheless, a rising awareness about benefits associated with apheresis is occurring. This awareness can ultimately lead toward early diagnosis of chronic disorders, consequently diluting the effect of the restraints.

North America is expected to have the largest market share owing to the highly sophisticated healthcare infrastructure and high healthcare expenditure. In addition, a high patient awareness, the demand for plasma-derived medicines, and the rise in the disposable income make this region dominant in the market. Asia-Pacific is also expected to grow at a high rate due to a considerably well-established apheresis market in Japan; moreover, emerging economies, such as India and China, have large patient pools, geriatric population, and rising healthcare expenditure.

Globally, the market depicts the presence of a highly intense competitive landscape due to the presence of innumerable vendors in the market, of which few hold a comparatively larger share in the market. Currently, new entrants in the market are not able to outshine their bigger rivals. However, the next few years are expected to change the scenario, as more companies start upping their game in the global apheresis market. Many businesses are focusing on strategies such as collaborations, mergers and acquisitions, and carrying out extensive research and development to grow in this market. In January 2018, MedAware Systems, Inc. launched a comprehensive database on apheresis through its SOHInfo division, carrying data from clinical trials, cohort studies, medical journals, and other sources.

The players are expected to tighten their grip by ferociously working toward gaining maximum revenue in the next few years for establishing themselves concretely in the global market. Kawasumi Laboratories Inc., B. Braun Melsungen AG, Terumo Corporation, Haemonetics Corporation, Medica S.p.A., Asahi Kasei Medical Co. Ltd., Therakos, Inc., HemaCare Corporation, Fresenius Kabi AG, Nikkiso Co., Ltd., and Cerus Corporation, are key companies operating in the global apheresis market since past few years.

Advances in blood bank centrifuges
Centrifuges are the backbone of a blood-banking lab. It is vital that the centrifuges should be reliable and able to provide fully reproducible and traceable data with every spin. With downstream patient-safety implications, separation of whole blood into its components needs to be tightly regulated and should be in compliance with GMP.

Centrifuges featuring large capacity in compact footprint, glove friendly center touch-interface, automatic door opening and closing with auto door, store lid functioning automatically with autolid are now available with remote monitoring and control systems. Password protection for multiuser environment, energy savings with eco-spin wind-shielded rotors, CFC-free refrigeration system with pre-cooling facility, maintenance free brushless induction motor, low-level of vibration, and noise with smooth acceleration and deceleration are some of the available upgrades of centrifuges for blood banks.

An option to open in case of power failure, easy loading and unloading facilities make them user friendly. Acceleration and deceleration time can be set as per user requirement. Provision of ACE function that corrects for variations in acceleration, centri-cross function that converts existing protocols, and connectivity for protocol tracking enrich the present day centrifuges. They are designed to meet production needs. Models accommodating large number and quantity of blood bags are at disposal in the market.

To a large extent, centrifuge providers have also eliminated a lot of the guesswork and streamlined their offerings through a combination of adaptability, ease of use, and vastly improved technology. For starters, providers are offering their main product ranges with all available rotors, attachments, and inserts under single part numbers to avoid costly and confusing a-la-carte shopping. With the exception of floor-model ultracentrifuges, units are typically designed to be plug-and-play, with little or no specialized knowledge required for immediate use.

Increased capacity and decreased negative space have reduced energy and size footprints, improving efficiency and savings over the long term. Additionally, material improvements to interchangeable parts have extended life spans and reduced the need for repair and maintenance contracts. For instance, the move from aluminum to carbon fiber rotors has diminished corrosion problems and allowed for longer warranties, and fast, foolproof rotor-swapping mechanisms have improved safety.

Advances in blood cell separators
FACS is one of the common technologies used in research environments for separation of cells and can isolate sub populations by tagging of multiple surface antigens. However, one of the main challenges with FACS is that the throughput and recovery is relatively low compared to other systems, and a typical run may be very time consuming for sorting of rare populations or large starting doses.

MACS that makes use of antibodies conjugated to magnetic beads is currently the most common method for isolating a population of cells, as it is quicker and has greater scalability than FACS. But it lacks the ability to release the magnetic beads from the isolated cell populations, which may be undesirable for immunotherapies prior to infusion into the patient. To overcome these challenges, a new technology that sorts blood cells using acoustic waves, referred to as acoustophoresis, is being developed. Benefits of using acoustic forces on micro scales for cell sorting include precise spatial control, fast action/switch rate, and little interference with cell viability. In microfluidic devices that use acoustic forces for sorting and manipulation, standing wave-type interaction is more popular.

Many types of microfluidic cell sorting devices have been reported recently to tackle the challenge of rare cell isolation from blood. External forces, including magnetic, electric, acoustic and optical, have been used in active microfluidic systems for focusing and extraction of target cells from suspensions. Meanwhile, passive systems that rely purely on channel geometry, carrier fluid, and cell properties have received attention due to their simplicity and high throughput. These include deterministic lateral displacement (DLD), pinched flow fractionation (PFF), hydrodynamic filtration, inertial migration, viscoelastic focusing, and their combinations.

Additionally, biological affinity has been widely used to target specific cell surface markers and improve selectivity of microfluidic cell sorting. While tremendous progress has been achieved, these platforms are not able to work with unprocessed whole blood and generally require a number of sample preparation steps, including lysis of red blood cells (RBCs), immunoselection, or sample dilution. Direct separation of cells from whole blood remains largely unexplored despite the persistent interest.

Lab-on-a-compact disk (LOCD) is also gaining much attention, due to notable advantages including cost-effective device fabrication, simple flow control by centrifugal force, compact instrumentation, and multi-processing on a single CD. The LOCD technique has been applied to blood analysis to determine hematocrit and blood cell separation. More recently, three-dimensional (3D) printing has been widely used due to its advantages such as low cost, easy access, various materials including biomaterials, and less restriction on design. Especially, 3D multi-layered microfluidic structures can be easily fabricated by 3D printing, compared to conventional soft lithography, which is limited by complex, multiple fabrication steps and precise alignment.

Novel technologies that are being developed and could be used in the future for cell separation include acoustic separation technology, which isolates cells of interest based on varied sizes of antigen–bead combinations in a closed, GMP-compliant manner; acoustic wave separation technology, capable of fluid–particle and fluid–fluid separation based on ultrasonic standing waves; and BACS which uses antibodies attached to microbubbles to isolate cells of interest. As knowledge of the properties of cells improves and tunable conjugation chemistry evolves, it is anticipated that greater control and specificity of antibody-based selection will also evolve, bringing costs down, and reducing processing time and complexity for the operator.

Outlook

Latest advances in cell biology, disease diagnostics, and medicine have increased the demand in rapid, safe, and accurate cell sorting and manipulation devices. Microfluidic devices are at the center of attention due to low sample and reagent volume requirement, portability, ability to work on a single cell scale level, and self-contained nature allowing safer handling of hazardous liquids and materials. Despite the advantages of performing cell analysis, sorting, and manipulation in a microfluidic chip, they still have a number of limitations that prevent standardization for clinical use and wide commercialization. Among these are device throughput, lifespan, multipart manufacturing, and ease of handling.

Multimodal, parallel integration of microfluidics with active sorting and manipulation methods is a promising approach to overcoming these limitations. Magnetic, electric, acoustic, and optical forces can be harnessed to cater to a wide spectrum of applications.

Moreover, optical forces can be applied from the outside of the microfluidic device, thus allowing development of highly modular, multi-purpose systems for cell sorting and manipulation. Optical forces offer more interaction freedom, which can be adjusted in real-time. Further investigation and development of novel techniques utilizing optical forces might prove to be a stepping stone toward development of state-of-the-art lab-on-a-chip devices.

Rapid technological advancements are leading to the development of next-generation apheresis devices with automated interface systems, screen navigation, and graphical user interface displays that will reduce human intervention and provide results faster and more effectively. Technology vendors – specifically digital companies – can play a larger role in enhancing apheresis product features.