單克隆抗體，那些你不知道的小秘密

Monoclonal antibodies，one of the most widely used tools in biological research, with the potential soon to become important therapeutics, have been with us for just over 40 years. These antibodies are monovalent, bind to a single epitope and are produced by a single B-lymphocyte clone. The first monoclonal antibody was generated in mice in 1975 by Milstein and Kohler, using the hybridoma technique. 

單克隆抗體是生物學(xué)研究中使用較廣泛的工具之一，也具有成為重要治療藥物的潛力，它的出現(xiàn)已有 40 多年的歷史。這些抗體是單價的，與單個表位結(jié)合并且由單個 B 淋巴細(xì)胞克隆產(chǎn)生。1975 年，Milstein 和 Kohler 使用雜交瘤技術(shù)在小鼠中產(chǎn)生了第一個單克隆抗體。

1.History and production 歷史和產(chǎn)生

Milstein and Kohler realised that in order to get an antibody where each molecule was identical to one another, they could harvest single B-cells from the spleen of a mouse and fuse them to myeloma (cancer) cell line, producing hybridomas.Antibody-producing B-cells are clonal, derived from memory B-cells, which means that every B-cell generated from a single memory B-cell produces the exact same antibody. These are termed ‘Monoclonal antibodies’ (single clone).

Milstein 和 Kohler 意識到，為了獲得每個分子彼此相同的抗體，他們從小鼠脾臟中收獲單個 B 細(xì)胞并將其融合到骨髓瘤（癌癥）細(xì)胞系中，從而產(chǎn)生雜交瘤。產(chǎn)生抗體的 B 細(xì)胞是來自記憶 B 細(xì)胞的克隆，這意味著從單個記憶 B 細(xì)胞產(chǎn)生的每個 B 細(xì)胞產(chǎn)生相同的抗體。這些被稱為「單克隆抗體」（單克?。?/strong>

Hybridoma production is still the most common method of Monoclonal antibody production. The fusions that are stable enough to grow in vitro are selected, sub-cloned and form a monoclonal-antibody producing hybridoma. These clones are then cultured and the antibodies they produce are purified either from the cells themselves, or the cell culture lysates. Each monoclonal antibody is defined by its unique binding site on the target protein, and this unique binding determines its specificity.

雜交瘤生產(chǎn)仍然是單克隆抗體生產(chǎn)中常用的方法。選擇足夠穩(wěn)定以在體外生長的融合體，亞克隆并形成產(chǎn)生單克隆抗體的雜交瘤。然后培養(yǎng)這些克隆，并將它們產(chǎn)生的抗體從細(xì)胞本身或細(xì)胞培養(yǎng)裂解物中純化。每種單克隆抗體由其在靶蛋白上的結(jié)合位點定義，并且這種的結(jié)合決定了其特異性。

 

2.Uses and alternative production methods 用途和替代生產(chǎn)方法

Antibodies with this level of specificity are a popular research tool for the detection of a single protein. They can also be used as a probe to locate the protein of interest inside a cell or organ. In recent years, alternative methods of producing monoclonal antibodies have become available. The so-called phage-display method meant that recombinant versions of the antigen-binding part of the antibody could be screened for specificity from a library of fusions to a carrier phage protein. Following on, further refinements to recombinant antibody production resulted in cell line-producing recombinant antibodies where the species of the constant region can be customized. The advantage of this new production method is that the issues around long-term stability of a hybridoma, with potential loss of a precious clone, is avoided.

The most important reason to use a monoclonal antibody has to be its consistency. As the cell-line is essentially immortal and unchanging, no new animals are needed for further production and the antibody remains consistent from batch to batch in its performance.

具有這種特異性水平的單克隆抗體是檢測單一蛋白質(zhì)中非常流行的研究工具。它們也可以用作探針來定位細(xì)胞或器官內(nèi)感興趣的蛋白質(zhì)。近年來，生產(chǎn)單克隆抗體的替代方法也已經(jīng)產(chǎn)生。如所謂的噬菌體展示方法，即從融合文庫篩選特異性抗體的抗原結(jié)合部分，以重組形式與噬菌體蛋白質(zhì)進(jìn)行融合表達(dá)。接下來，對重組抗體生產(chǎn)的進(jìn)一步改進(jìn)而產(chǎn)生細(xì)胞系的重組抗體，其中恒定區(qū)的種類可以定制。這種新的生產(chǎn)方法的優(yōu)點是克服了雜交瘤長期不穩(wěn)定性可能會損失寶貴的克隆的問題。

使用單克隆抗體最重要的原因是它的一致性。由于細(xì)胞系基本上是不朽且不變的，所以不需要新的動物用于進(jìn)一步生產(chǎn)，并且抗體在其性能中從批次到批次保持一致。
 

In addition to the use of the entire immunoglobulin, for certain experiments these immunoglobulin molecules can be cleaved to give smaller versions with specific properties. The the constant region (Fc) of the immunoglobulin can be cleaved either above or below the hinge region, leaving either (Fab’)2 or Fab fragments. Since the Fc portion has affinity to Ig receptors, the removal of this part of the antibody reduces non-specific binding and is therefore often used in Flow cytometry applications or where binding to Fc receptors must be avoided.

除了使用完整的免疫球蛋白之外，對于某些實驗，這些免疫球蛋白分子可以被切割以產(chǎn)生具有特定性質(zhì)的片段。免疫球蛋白的恒定區(qū)（Fc）可以在鉸鏈區(qū)上方或下方切割，留下（Fab'）2 或 Fab 片段。由于 Fc 部分對 Ig 受體具有親和力，所以抗體這部分的去除減少了非特異性結(jié)合，因此經(jīng)常這種抗體常用于流式細(xì)胞術(shù)應(yīng)用或者其他必須避免結(jié)合 Fc 受體的免疫實驗。

Biorbyt 擁有的人，小鼠，大鼠，兔各種來源的單克隆抗體，優(yōu)勢和克隆統(tǒng)計數(shù)據(jù)如下。感興趣的請聯(lián)系我們，獲取更多單克隆抗體的產(chǎn)品信息和咨詢。
 

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