B Cell Immunophenotyping

1.The background of B cells

B lymphocytes, abbreviated as B cells, are differentiated from hematopoietic stem cells in the bone marrow. Most of the B cells that exert effector function live in the body for a short period of time, only a few days to several weeks, but their memory cells can exist for a long time in the body.

B cells are also heterogeneous that exhibit different functions and surface markers. Whether CD5 is expressed or not, B cells are divided into two subsets of B1 and B2. B1 cells are CD5+ B cells, which mainly recognize non-protein antigens and belong to intrinsic lymphocytes. B2 is usually referred to as B cells, which are CD5-B cells and mainly recognize protein antigens. With the help of Th cells, B2 cells can be fully activated and mediate T cell-dependent antigen-dependent immune responses, producing specific antibodies. The B2 cell-mediated immune response is characterized by somatic mutations, affinity maturation, produces high-affinity antibodies, and immunological memory cells.

2.B cell differentiation and maturation
The differentiation and maturation of mammalian B cells are mainly divided into several stages: progenitor B cells, pre-B cells, immature B cells, B cell maturation (transition B cells to mature B cells), activated B cells and plasma cells.

Progenitor B cell (pro-B cell)
This early development of B cells occurs around the 9th week of human embryos and begins on day 14 of the mouse. The specific surface markers of the B cell line have not been expressed, and no lg gene rearrangement has occurred, and it is still in the germline stage. However, late B-cells may have B-specific markers, such as Thy-1, Tdt, B200, and mb-1.

Pre-B cell
The pre-B cells are differentiated from progenitor B cells in the bone marrow, and are present only in hematopoietic tissues such as bone marrow and fetal liver, accounting for about 5% of adult bone marrow nucleated cells. The heavy chain μ of IgM can be detected in the cytoplasm of pre-B cells, but no light chain, and no expression of Ig on the membrane surface, thus lacking the ability to respond to antigens.

Immature B cell
At this stage, light chain gene rearrangement occurs, so that a complete IgM molecule can be formed and expressed on the surface of the membrane (IgM), which can be called Bμcells. Immature B cells begin to lose Tdt and CD10, but express CD22, CD21 and FcR. At the same time, the expression levels of CD19, CD20 and MHC class II molecules increase.

B cell maturation
Immature B cells will leave the bone marrow within a few days and enter the circulatory system as transitional B cells. Transitional B cell maturation occurs mainly in the spleen, and T1 B cells are matured into T2 B cells by ligand stimulation of the BAFF family and receptor-mediated signaling.

Activated B cells

Mature B cells are stimulated by the corresponding antigens or polyclonal stimulators to become activated B cells, which then proliferate and differentiate. During this process, the level of membrane-bound Ig gradually decreases, while the secretory Ig gradually increases, and the conversion of the immunoglobulin gene heavy chain class occurs. A part of activated B cells can differentiate into small lymphocytes, stop proliferation and differentiation, and can survive for several months to several years. When contacted with the same antigen again, the antibody’s activation and differentiation occur rapidly, and it has a short incubation period and antibody level. High and long-lasting, this B cell is called memory B cell.

Plasma cells (PC)
It is also known as antibody secreting cell. After the mature B cells are stimulated by the antigen, they become activated B cells with the help of antigen-presenting cells and Th cells, and then differentiate into plasma cells. When mature B cells differentiate into plasma cells, some of the markers on the surface of B cells disappear, and some new plasma cell-specific markers, such as plasma cell antigen-1 (PCA-1), are present, while mIg, MHC class II antigen, CD19 Marks such as CD20, CD21, and CD22 disappear.