抗體的生物素化標記實驗要點：

1. 絲氨酸/蘇氨酸蛋白激酶PIM3抗體品牌 如在反應(yīng)混合液中有疊氮鈉或游離氨基存在,會抑制標記反應(yīng)。因此,蛋白質(zhì)在反應(yīng)前要對 0.1mol/L碳酸氫鈉緩沖液或0.5mol/L硼酸緩沖液充分透析；

2.所用的NHSB及待生物素化蛋白質(zhì)之間的分子比按蛋白質(zhì)表面的ε-氨基的密度會有所不同,選擇不當則影響標記的效率,應(yīng)先用幾個不同的分子比來篩選最適條件；

3.用NHSB量過量也是不利的,抗原的結(jié)合位點可能因此被封閉,導(dǎo)致抗體失活；

4.由于抗體的氨基不易接近可能造成生物素化不足,此時可加入去污劑如 Triton x-100, Tween20等；

5.當游離ε-氨基(賴氨酸殘基的氨基)存在于抗體的抗原結(jié)合位點時,或位于酶的催化位點時,生物素化會降低或損傷抗體蛋白的結(jié)合力或活性；

6.生物素還可能與不同的功能基團,如羰基、氨基、巰基、異咪唑基及苯酚基,也可與糖基共價結(jié)合；

7.交聯(lián)反應(yīng)后,應(yīng)充分透析,否則,殘余的生物素會對生物素化抗體與親和素的結(jié)合產(chǎn)生競爭作用；

8.在細胞的熒光標記實驗中,中和親和素的本底低,但由于鏈霉親和素含有少量正電荷,故對某些細胞可導(dǎo)致高本底。

產(chǎn)品訂購信息：
英文名稱  Anti-PIM3 

中文名稱   絲氨酸/蘇氨酸蛋白激酶PIM3抗體品牌 

別    名  Kid 1; Kid-1; Kid1; Kinase induced by depolarization; ONCOGENE PIM 3; ONCOGENE PIM3; PIM 3; Pim 3 oncogene; PIM3; Pim3 oncogene; PIM3_HUMAN; Protein kinase Kid 1; Protein kinase Kid1; Serine/threonine kinase Pim 3; Serine/threonine kinase Pim3; Serine/threonine protein kinase Pim 3; Serine/threonine protein kinase Pim3; Serine/threonine-protein kinase Pim-3.

濃    度   1mg/1ml

規(guī) 格  0.2ml/200μg

抗體來源   Rabbit

克隆類型   polyclonal

交叉反應(yīng)   Human, Mouse, Rat, Chicken, Dog, Pig, Cow, Rabbit, Sheep, Guinea Pig 

產(chǎn)品類型   一抗  

研究領(lǐng)域     細胞生物 信號轉(zhuǎn)導(dǎo) 激酶和磷酸酶

蛋白分子量  predicted molecular weight: 36kDa

性    狀   Lyophilized or Liquid

免 疫 原 KLH conjugated synthetic peptide derived from human PIM3 

亞    型   IgG

純化方法   affinity purified by Protein A

儲 存 液   Preservative: 15mM Sodium Azide, Constituents: 1% BSA, 0.01M PBS, pH 7.4

絲氨酸/蘇氨酸蛋白激酶PIM3抗體品牌 產(chǎn)品應(yīng)用    WB=1:100-500 ELISA=1:500-1000 IHC-P=1:100-500 IHC-F=1:100-500 IF=1:50-200

（石蠟切片需做抗原修復(fù)） 

 not yet tested in other applications.

 optimal dilutions/concentrations should be determined by the end user.  

保存條件  Store at -20 °C for one year. Avoid repeated freeze/thaw cycles. The lyophilized antibody is stable at room temperature for at least one month and for greater than a year when kept at -20°C. When reconstituted in sterile pH 7.4 0.01M PBS or diluent of antibody the antibody is stable for at least two weeks at 2-4 °C. 

Important Note  This product as supplied is intended for research use only, not for use in human, therapeutic or diagnostic applications. 

產(chǎn)品介紹 The Pim family serine/threonine protein kinases were first identified in studies examining genes targeted for proviral insertion in murine leukemia virus-induced T lymphomas. Increased levels of Pim kinases predispose cells to lymphomagenesis and enhance the activity of mitogenic proteins such as p100, c-Myb, and Cdc25A. In addition, Pim kinases are also involved in modulation of synaptic strength in neurons and anti-apoptotic signaling in hematopoietic progenitor cells. Pim-3, a member of the proto-oncogene Pim family that expresses serine/threonine kinase activity, shares significant homology with Pim-1 serine/threonine protein kinases. Pim-3 may function as a mediator of synaptic plasticity in the brain and is presumably involved in the anti-apoptosis process and cell cycle progression as well as the proliferation of human hepatoma cell lines. The Pim-3 protein is widely expressed, however no expression is observed in the colon, thymus, or small intestine.

Function : Proto-oncogene with serine/threonine kinase activity that can prevent apoptosis, promote cell survival and protein translation. May contribute to tumorigenesis through: the delivery of survival signaling through phosphorylation of BAD which induces release of the anti-apoptotic protein Bcl-X(L), the regulation of cell cycle progression, protein synthesis and by regulation of MYC transcriptional activity. Additionally to this role on tumorigenesis, can also negatively regulate insulin secretion by inhibiting the activation of MAPK1/3 (ERK1/2), through SOCS6. Involved also in the control of energy metabolism and regulation of AMPK activity in modulating MYC and PPARGC1A protein levels and cell growth.

Subunit : Interacts with BAD. Interacts with PPP2CA; this interaction promotes dephosphorylation of PIM3, ubiquitination and proteasomal degradation. Interacts with SOCS6 (By similarity).

Subcellular Location : Cytoplasm.

Post-translational modifications : Ubiquitinated, leading to proteasomal degradation.

Phosphorylated. Interaction with PPP2CA promotes dephosphorylation.

Similarity : Belongs to the protein kinase superfamily. CAMK Ser/Thr protein kinase family. PIM subfamily.

Contains 1 protein kinase domain.

Database links :

UniProtKB/Swiss-Prot: Q86V86.3

Entrez Gene: 415116 Human

Entrez Gene: 223775 Mouse

Omim: 610580 Human

SwissProt: Q86V86 Human

SwissProt: P58750 Mouse

Unigene: 530381 Human

Unigene: Hs.530381 Human

Unigene: 400129 Mouse

抗體的鑒定：

1）絲氨酸/蘇氨酸蛋白激酶PIM3抗體品牌 抗體的效價鑒定：不管是用于診斷還是用于,制備抗體的目的都是要求較高效價。不同的抗原制備的抗體,要求的效價不一。鑒定效價的方法很多,包括有試管凝集反應(yīng),瓊脂擴散試驗,酶聯(lián)免疫吸附試驗等。常用的抗原所制備的抗體一般都有約成的鑒定效價的方法,以資比較。如制備抗抗體的效價,一般就采用瓊脂擴散試驗來鑒定。

2）抗體的特異性鑒定：抗體的特異性是指與相應(yīng)抗原或近似抗原物質(zhì)的識別能力?？贵w的特異性高,它的識別能力就強。衡量特異性通常以交叉反應(yīng)率來表示。交叉反應(yīng)率可用競爭抑制試驗測定。以不同濃度抗原和近似抗原分別做競爭抑制曲線,計算各自的結(jié)合率,求出各自在IC50時的濃度,并按公式計算交叉反應(yīng)率。 

如果所用抗原濃度IC50濃度為pg/管,而一些近似抗原物質(zhì)的IC50濃度幾乎是無窮大時,表示這一抗血清與其他抗原物質(zhì)的交叉反應(yīng)率近似為0,即該血清的特異性較好。

3）抗體親和力：是指抗體和抗原結(jié)合的牢固程度。親和力的高低是由抗原分子的大小,抗體分子的結(jié)合位點與抗原決定簇之間立體構(gòu)型的合適度決定的。有助于維持抗原抗體復(fù)合物穩(wěn)定的分子間力有氫鍵,疏水鍵,側(cè)鏈相反電荷基因的庫侖力,范德華力和空間斥力。親和力常以親和常數(shù)K表示,K的單位是L/mol?？贵w親和力的測定對抗體的篩選,確定抗體的用途,驗證抗體的均一性等均有重要意義。

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