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Background and objectives: Paper biodiagnostics for blood typing are novel, cheap, fast and easy to use. Agglutinated red blood cells cannot travel through the porous structure of paper, indicating a positive antibody-antigen interaction has occurred. Conversely, non-agglutinated blood can disperse and wick through the paper structure with the ease to indicate a negative result. This principle has been demonstrated to detect blood group phenotypes: ABO and RhD. However, typing for red blood cell antigens such as Rh, Kell, Duffy and Kidd has not yet been explored on paper.

Materials and methods: Two paper testing methods - an elution and a direct flow-through method - were investigated to detect red blood cell antigens excluding the ABO system and RhD. Antigens explored include the following: C, c, E, e, K, k, Fy(a), Fy(b), Jk(a), Jk(b), M, N, S and s, P1, Le(a) and Le(b). The variables tested include the following: reaction time and reagent concentration. The importance of antibody type/structure for successful agglutination on paper was confirmed.

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Results: Some blood group phenotypes showed less agglutination due to weaker antibody-antigen interactions. Most blood groups with antibodies available as IgM, such as C, c, E, e, K and k, and Jk(a) and Jk(b), and P1, were successful using both methods. However, other blood groups, especially those with antibodies only available as polyclonal antibodies, were unsuccessful and require further scrutiny.

Blood typing is a method to tell what type of blood you have. Blood typing is done so you can safely donate your blood or receive a blood transfusion. It is also done to see if you have a substance called Rh factor on the surface of your red blood cells.

Your blood type is based on whether or not certain proteins are on your red blood cells. These proteins are called antigens. Your blood type (or blood group) depends on what types your parents passed down to you.

A blood sample is needed. The test to determine your blood group is called ABO typing. Your blood sample is mixed with antibodies against type A and B blood. Then, the sample is checked to see whether or not the blood cells stick together. If blood cells stick together, it means the blood reacted with one of the antibodies.

The second step is called back typing. The liquid part of your blood without cells (serum) is mixed with blood that is known to be type A and type B. People with type A blood have anti-B antibodies. People with type B blood have anti-A antibodies. Type O blood contains both types of antibodies.

There is little risk involved with having your blood taken. Veins and arteries vary in size from one person to another, and from one side of the body to the other. Taking blood from some people may be more difficult than from others.

There are many antigens besides the major ones (A, B, and Rh). Many minor ones are not routinely detected during blood typing. If they are not detected, you may still have a reaction when receiving certain types of blood, even if the A, B, and Rh antigens are matched.

Westhoff CM, Storry JR, Shaz BH. Human blood group antigens and antibodies. In: Hoffman R, Benz EJ, Silberstein LE, et al, eds. Hematology: Basic Principles and Practice. 7th ed. Philadelphia, PA: Elsevier; 2018:chap 110.

Another microfluidic device was developed to type the ABO/RH(D) blood groups and identify the weak antigens (49, 50). The process requires only 1l of blood collected by pricking the fingertip, which is then mixed with 10 l of PBS in a reaction tank (49). The sample passes through a four-layer microfluidics chip pre-loaded with anti-A, anti-B, and anti-D antibodies (0.5 l). The agglutinated RBCs accumulate in the channel, and the formation of a distinct red line within 2 min is a positive result. The length of the red line is directly proportional to the agglutination intensity. A microfluidic thread-based analytical device was also used to accurately identify six weak A subgroups and 89 normal ABO groups (50). This device can use a manual pump system that does not need to rely on electricity and therefore can be used outdoors, at home, or in an emergency vehicle (51).

However, the aforementioned techniques generally require DNA purification, which is usually the most speed-limiting and labor-intensive step. On the other hand, direct, real-time allele-specific PCR and melting curve analysis (DRAM) is a fast and reliable one-step blood group genotyping technique that does not require DNA preparation (122). It uses a special buffer for direct PCR, rapid RBC lysis buffer, white blood cell DNA template, allele-specific primers, and DNA-binding fluorescent dyes (EvaGreen; Biotium) for discriminating ABO alleles. The PCR reaction process is carried out in a closed system, reducing manpower and material resources and the risk of contamination. Studies showed that DRAM measurement was 100% consistent with the ABO genotyping results of PCR-RFLP, PCR direct sequencing, and serological typing results. Compared to traditional ABO genotyping utilizing allele-specific PCR with purified DNA and agarose gel electrophoresis, DRAM reduces manual procedures to hands-on time from ~40 to 12 min. The total time required for the DRAM assay is around 274 min.

Additionally, NGS has the capacity to sequence the whole genome (WGS), but is neither cost-efficient nor practical for clinical transfusion laboratories. Nevertheless, WGS is increasingly performed to discover unknown or undiscovered allelic polymorphisms, blood group antigen phenotypes, and associations between genes and certain chronic diseases (105). In one study (123), the whole genome data of 79 individuals with nine red blood cell antigen systems was analyzed, and the consistency of blood group polymorphism was 93%. A total of 267 gene polymorphisms identified in this study were not present in the ISBT database. The highly complex ABO/RH(D) and MNS systems were also identified by WGS. Blood group typing with WGS is feasible but requires improvements in reading depth for precision typing of single nucleotide variant polymorphisms (123). An automated RBC typing algorithm based on WGS data was 100 % concordant with serological methods for ABO and D antigens and 99.5% accurate at typing the C antigen (124). In the future, WGS may play an essential role in detecting rare blood types and RBC diseases.

Indeed, pre-transfusion compatibility testing largely relies on serological testing, but antibody reagents are not available for a large number of rare blood group antigens testing with regard to rare blood requirements. Genotyping overcomes serology limitations, has the capacity of high-throughput testing, and allows easier detection of rare blood group antigens. As already discussed, PCR-RFLP, PCR-SSCP, DNA sequencing, etc., are the common genetic tests used for blood typing. They can be used to identify rare blood types and study the inheritance and frequency of rare blood antigen genes, such as real-time fluorescent PCR (95, 96). NGS is also suitable for identifying rare blood types without knowing specific antibodies (110, 111). Currently, there are more rapid and cost-effective genotyping kits with multiplex capacity and high-throughput volumes, enabling affordable large-scale rare blood group genotyping (127).

The need to first zero in on a blood group can delay blood transfusions in emergency situations, and this in turn can prove fatal. Thus, to speed up the process, a team of scientists from Tokyo University of Science, Japan, has developed a lab-on-a-chip device that can not only tell the blood type within five minutes but allows medical staff to read the results through simple visual inspections.

Blood transfusion, if performed promptly, is a potentially life-saving intervention for someone losing a lot of blood. However, blood comes in several types, some of which are incompatible with others. Transfusing an incompatible blood type can severely harm a patient. It is, therefore, critical for medical staff to know a patient's blood type before they perform a transfusion.

There are four major blood types--O, A, B, and AB. These types differ based on the presence or absence of structures called A antigens and B antigens on the surfaces of red blood cells. Blood can be further divided into positive and negative types based on the presence or absence of D antigens on red blood cells. Medical professionals usually tell a patient's blood type with tests involving antibodies against the A and B antigens. When antibodies recognize the corresponding antigens, they bind to them, causing the blood cells to clump together and the blood to coagulate. Thus, specific antigen-antibody combinations tell us what the blood type of a blood sample is.

Aiming to solve these problems, a team of scientists at Japan's Tokyo University of Science, led by Dr Ken Yamamoto and Dr Masahiro Motosuke, has developed a fully automated chip that can quickly and reliably determine a patient's blood type. In the words of Dr Motosuke, he and his colleagues "have developed a compact and rapid blood-typing chip which also dilutes whole blood automatically."

The chip contains a micro-sized "laboratory" with various compartments through which the blood sample travels in sequence and is processed until results are obtained. To start the process, a user simply inserts a small amount of blood, presses a button, and waits for the result. Inside the chip, the blood is first diluted with a saline solution and air bubbles are introduced to promote mixing. The diluted blood is transported to a homogenizer where further mixing, driven by more intensely moving bubbles, yields a uniform solution. Portions of the homogenized blood solution are introduced into four different detector chambers. Two chambers each contain reagents that can detect either A antigens or B antigens. A third chamber contains reagents that detect D antigens and a fourth chamber contains only saline solution, with no reagent, and serves as a negative control chamber in which the user should not observe any results. Antigen-antibody reaction will cause blood to coagulate, and by looking at which chambers have coagulated blood, the user can tell the blood type and whether the blood is positive or negative. 006ab0faaa