培養方法

傳代方法 將舊培養液吸除，PBS清洗兩遍后，加入6mL（/100mm皿）胰酶，在顯微鏡下觀察，期間禁止搖晃培養皿，細胞剛有脫落時，則吸除大部分胰酶，留約0.5mL，移至培養箱消化，約2min取出。傳代用12mL CM1-1培養液終止消化，輕輕吹打均勻細胞，后可分3~6皿培養；

生長條件 37℃，5%CO2，CM1-1培養液。CM1-1培養液：90%DMEM-H+10%FBS。DMEM-H：DMEM高糖培養液，含谷氨酰胺，含丙酮酸鈉。

存儲條件 凍存則用6mL凍存液（90%FBS+10%DMSO）終止消化，吹打均勻，分為6支凍存管，用程序降溫盒于-80℃凍存，過夜轉移至液氮中保存。

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培養操作步驟 ：

1．用蓋片鑷將蓋玻片自75%乙醇中取出，用無菌絲綢布擦拭干凈，不要用紗布； 

2．將蓋玻片輕輕放入6孔培養板（每孔一片）或培養皿中（每個平皿可放置2-3片）； 

3．在距離紫外燈直射范圍內20-30 厘米處照射2-3小時； 

4．將經過計數的細胞懸浮液移入培養板中，使蓋玻片完全浸在培養液中； 

5．將培養板在5% CO2水浴孵箱中37℃孵育2-3天，當貼壁細胞生長至覆蓋培養板底部2/3面積時，將培養板取出，用蓋片鑷輕輕取出蓋玻片，用蒸餾水漂洗后即可進行快速固定以及細胞化學檢測。 

細胞培養方法：

1、細胞傳代：細胞密度達到80-90%時即可傳代

①棄去培養上清，用PBS或生理鹽水清洗1-2次；

②加入2ml0.25%胰酶（T25瓶），使胰酶覆蓋整個瓶或皿，蓋好放入培養箱消化；

③1-2min后，顯微鏡下觀察細胞，若大部分細胞回縮且有少量細胞脫落，輕輕吹打下確認消化情況后加入完全培養基終止消化；若細胞還是貼壁，放回培養箱繼續消化至可以輕輕吹打下為止；

④將細胞懸液1000RPM左右條件下離心4min，棄上清；

⑤用新鮮培養基重懸后加入培養瓶或皿中，T25培養瓶加6-8ml培養基；

⑥懸浮細胞直接離心收集，細胞沉淀重懸后分到新培養瓶中。

2、細胞復蘇：

①將凍存管在37℃溫水中快速搖晃融化，時間1min左右，加入4-5ml培養基混勻。

②在1000RPM左右條件下離心4min，棄上清,加1-2ml培養基吹勻，將細胞懸液加入培養瓶中，補加適量培養基。

3、細胞凍存：待細胞生長狀態良好時進行細胞凍存保種

①棄去培養上清，用PBS或生理鹽水清洗1-2次，加入1mL 0.25%胰蛋白酶（T25瓶）

②1-2min后，顯微鏡下觀察細胞，大部分細胞回縮且有少量細胞脫落，輕輕吹打下確認消化情況后加入完全培養基終止消化；

③將細胞懸液1000RPM左右條件下離心4min，棄上清，加1ml凍存液重懸細胞；

④將凍存管放入程序降溫盒，放入-80℃冰箱，4小時后將凍存管轉入液氮罐儲存。

（石蠟切片需做抗原修復）

 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.

產品介紹 Otoancorin, also known as OTOA, CT108 or DFNB22, is a 1,153 amino acid protein belonging to the stereocilin family. Expressed in the inner ear and restricted to the interface between the apical surface of sensory epithelia, otoancorin is suggested to act as an adhesion molecule. Otoancorin ensures the attachment of the inner ear acellular gels to the apical surface of the underlying nonsensory cells. Mutations in the gene encoding otoancorin leads to deafness autosomal recessive type 22 (DFNB22), which is a form of sensorineural hearing loss. Sensorineural deafness results from damage to the neural receptors of the inner ear, the nerve pathways to the brain or the area of the brain that receives sound information. Existing as three alternatively spliced isoforms, otoancorin is encoded by a gene located on human chromosome 16p12.2.

Function : May act as an adhesion molecule.

Subcellular Location : Apical cell membrane. Secreted > extracellular space > extracellular matrix. At the interface between the apical surface of the epithelia and the overlying acellular gel of the tectorial and otoconial membranes.

DISEASE : Defects in OTOA are the cause of deafness autosomal recessive type 22 (DFNB22) [MIM:607039]. DFNB22 is a form of sensorineural hearing loss. Sensorineural deafness results from damage to the neural receptors of the inner ear, the nerve pathways to the brain, or the area of the brain that receives sound information.

Similarity : Belongs to the stereocilin family.

Database links : UniProtKB/Swiss-Prot: Q7RTW8.1

英文名稱  Anti-Osteocalcin

中文名稱  骨鈣蛋白/骨鈣素抗體

別    名  Osteocalcin Propeptide; Osteocalcin; Osteocalcin precursor; Gamma-carboxyglutamic acid-containing protein; Bone Gla-protein; BGLAP; Bone gamma carboxyglutamate protein; BGLAP; BGP; Bone gamma carboxyglutamate gla protein osteocalcin; Bone gamma carboxyglutamate protein; Bone Gla protein; Gamma carboxyglutamic acid containing protein; OC; PMF1; OSTCN_HUMAN.

濃    度  1mg/1ml

規 格  0.1ml/100μg  0.2ml/200μg  1ml/1mg

抗體來源  Rabbit  

克隆類型  polyclonal
小鼠結腸癌細胞熒光素酶標記骨橋素檢測試劑盒脫氫乙酸鈉 99%DL-乳酸 ACS, ≥85%

骨特異性堿性磷酸酶檢測試劑盒脫氫乙酸 98%L-乳酸鈉  AR，60%水溶液

骨特異性堿性磷酸酶B檢測試劑盒乙酰乙酸乙酯  >99.0%(GC)L-乳酸鈉 98%

骨退化特異標志物檢測試劑盒乙酰乙酸甲酯  99%6-氨基吲唑 98%

骨涎蛋白檢測試劑盒3-基吡啶 99%間羥基苯甲酸 98%

瓜氨酸檢測試劑盒鄰苯二甲 98%5-硝基吲唑 98%

寡克隆區帶檢測試劑盒山梨酸 >99.0%(T)6-硝基吲唑 98%

實驗要點及說明：

1．本方法適用于貼壁細胞培養，而不適用于懸浮細胞培養，懸浮細胞可使用滴片法； 

2．所使用的蓋玻片應該為玻璃**，并經過鉻酸洗液處理； 

3．蓋玻片非常薄，易碎，取放蓋玻片時動作要輕； 

4．如果需要更多生長狀態一致的細胞，可以使用較大的培養皿，但不宜過大，以避免培養液的浪費和增加污染機率； 

5．如果細胞貼壁生長能力較差，可將蓋玻片在0.5%多聚賴氨酸溶液中浸泡5-10分鐘并自然晾干。