臺灣博碩士論文加值系統

從台灣長牡蠣的外殼珍珠質中，萃取出多種的蛋白質分子，並且對這些被萃取出的蛋白質分子進行初步的生化特性分析與生物活性測試。廣佈於文石內的蛋白質，在經過醋酸的去礦物質化後，部分可溶解於醋酸溶液中，此部分萃取物稱之為酸萃取物；而因聚集作用而不溶解的或吸附在不溶解於醋酸之基質上的蛋白質，則使用十二基硫酸鈉聚丙烯醯胺與乙基硫醇將之萃取出來，此部分萃取物稱之為乙基硫醇萃取物。此兩種萃取物在經過膠體電泳，將所含的蛋白質分離之後，利用多種的染色法來測定所含蛋白質的種類與特性；於長牡蠣珍珠質的乙基硫醇萃取物中，發現有四種醣蛋白的存在；利用Stains-all與Alcian blue染色法，偵測出在乙基硫醇萃取物內含有兩種多陰離子性蛋白質，可能與鈣離子的結合有關。令人好奇的是，在分子量45與40kDa的位置，同時於醣蛋白與多陰離子性蛋白質的染色皆呈現陽性結果，而這兩個位置上的蛋白質是否為同一種蛋白，或只是剛好分子量相似的兩種相異蛋白，則需要再作進一步的分析。U2-OS是一種類造骨細胞株，能在體外培養時進行自發性的礦物質化，利用U2-OS具有造骨細胞的特性，將細胞與萃取物共同培養7天，藉以觀察酸萃取物與乙基硫醇萃取物是否具有生物活性；在細胞型態觀察方面，乙基硫醇萃取物在第7天時，能明顯的促進細胞產生鈣化現象，而酸萃取物則沒有影響；在活化細胞鹼性磷酸酶活性方面，不管是酸萃取物或是乙基硫醇萃取物，皆無法促進鹼性磷酸酶活性的增加。總而言之，在台灣長牡蠣的珍珠質中，不但發現有醣蛋白以及多陰離子性蛋白質的存在，更發現珍珠質的乙基硫醇萃取物具有促進細胞鈣化的生物活性。

目錄-------------------------------------------- Ⅰ
圖次-------------------------------------------- Ⅲ
中文摘要---------------------------------------- 1
英文摘要---------------------------------------- 2
壹、前言---------------------------------------- 4
貳、文獻回顧------------------------------------ 6
2.1 珍珠層----------------------------------- 6
2.2 水溶性基質------------------------------- 9
2.3非水溶性基質------------------------------ 13
2.4體內與體外研究---------------------------- 15
2.5骨的形成---------------------------------- 17
2.6鹼性磷酸酶-------------------------------- 18
參、材料與方法----------------------------------- 20
一、實驗材料---------------------------------- 20
1. 藥品及試劑------------------------------- 20
2. 儀器設備--------------------------------- 21
二、細胞株資料單------------------------------ 22
三、實驗方法---------------------------------- 24
3.1珍珠層粉末（nacre power）來源------------- 24
3.2樣本製備--------------------------------- 24
3.3 SDS-polyacrylamide gel electrophoresis (SDS-PAGE)-------------------------------------- 26
3.4硝酸銀染色（silver stain）------------------ 30
3.5 醣蛋白測定------------------------------ 31
3.6 Alcian blue 染色法------------------------ 31
3.7 Stains-all 染色法-------------------------- 32
3.8細胞培養--------------------------------- 32
3.9 Von-Kossa 染色法------------------------- 33
3.10 鹼性磷酸酶活性測定--------------------- 33
肆、結果----------------------------------------- 34
伍、討論----------------------------------------- 37
陸、結論----------------------------------------- 41
參考文獻----------------------------------------- 49

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