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ADPG焦磷酸化酶(AGP)活性檢測試劑盒

操作步驟：

1,分光光度計預熱 30min,調(diao)節波長到 340nm,蒸餾水調(diao)零(ling)。并將試劑一置(zhi) 30℃保溫 10min 以上。

2，在(zai)離心管(guan)中按(an)順序加入(ru)以(yi)下(xia)試(shi)(shi)(shi)劑(ji)(ji)(ji)：測定管(guan)試(shi)(shi)(shi)劑(ji)(ji)(ji)一（μL） 100 試(shi)(shi)(shi)劑(ji)(ji)(ji)二（μL） 10 試(shi)(shi)(shi)劑(ji)(ji)(ji)三(san)（μL） 50 試(shi)(shi)(shi)劑(ji)(ji)(ji)四（μL） 100 樣(yang)本（μL） 20 混勻，30℃保(bao)溫(wen) 15 min，置沸(fei)水浴中 1 min，冰浴迅速冷(leng)卻試(shi)(shi)(shi)劑(ji)(ji)(ji)一（μL） 300 試(shi)(shi)(shi)劑(ji)(ji)(ji)五（μL） 100 試(shi)(shi)(shi)劑(ji)(ji)(ji)六（μL） 10 試(shi)(shi)(shi)劑(ji)(ji)(ji)七（μL） 10 混勻后立(li)即在(zai) 340 nm 波長(chang)下(xia)記(ji)錄初始吸(xi)光(guang)度 A1 和 2min 后的吸(xi)光(guang)度 A2，計算ΔA=A2-A1。

ADPG焦磷酸化酶(AGP)活性檢測試劑盒

AGP 活性計算：

1、按樣本蛋白濃度計算：單位(wei)的定義：每 mg 組(zu)織(zhi)蛋白在反應體系中(zhong)每分鐘催(cui)化(hua)產生 1nmol NADPH 定義為(wei)一個酶活性單位(wei)。 AGP（U/mg prot）＝[ΔA×V 反總÷(ε×d)×10 9] ÷（V 樣×;Cpr）÷T=2813×ΔA÷Cpr。此(ci)法需要自行測定粗酶液(ye)蛋白質濃度。

2、按照樣本鮮重(zhong)計算(suan)單位的定義：每 g 組(zu)織在(zai)反(fan)應體系中每分鐘(zhong)催(cui)化產生 1nmol NADPH 定義為(wei)一(yi)個酶活力單位。 AGP 活性(xing)（U/g mass）=[ΔA×V 反(fan)總(zong)÷(ε×d)×10 9] ÷（W×V 樣÷V 樣總(zong)）÷T=2813×ΔA÷W

V 反總(zong)(zong)：反應(ying)體系總(zong)(zong)體積(ji)，7×10 -4 L；ε：NADPH 摩爾(er)消光系數(shu)，6.22×10 3mol/L/cm；V 樣：加入(ru)樣本體積(ji)，0.02 mL；

V 樣總：加入提取液體積(ji)，1 mL；d:比(bi)色皿(min)光徑 1cm。Cpr：樣本(ben)蛋白質濃度，mg/mL。W：樣品質量，g。T:反應(ying)時間 2min。

ADPG焦磷酸化酶(AGP)活性檢測試劑盒

提取液(ye)：液(ye)體 30mL×1 瓶(ping)(ping)(ping)，4℃保(bao)(bao)(bao)存(cun)(cun); 試(shi)(shi)(shi)(shi)(shi)劑(ji)一：液(ye)體 10mL×1 瓶(ping)(ping)(ping)，4℃保(bao)(bao)(bao)存(cun)(cun)；試(shi)(shi)(shi)(shi)(shi)劑(ji)二(er)：粉(fen)劑(ji)×1 支(zhi)(zhi)，4℃保(bao)(bao)(bao)存(cun)(cun)；臨(lin)(lin)(lin)用(yong)(yong)(yong)(yong)前(qian)(qian)(qian)每(mei)支(zhi)(zhi)加(jia)入 250μL 雙(shuang)(shuang)蒸(zheng)水(shui)充分(fen)(fen)溶(rong)解(jie)備用(yong)(yong)(yong)(yong)；用(yong)(yong)(yong)(yong)不(bu)完的(de)試(shi)(shi)(shi)(shi)(shi)劑(ji)仍(reng) 4℃保(bao)(bao)(bao)存(cun)(cun)；試(shi)(shi)(shi)(shi)(shi)劑(ji)三：粉(fen)劑(ji)×1 瓶(ping)(ping)(ping)，4℃保(bao)(bao)(bao)存(cun)(cun)；臨(lin)(lin)(lin)用(yong)(yong)(yong)(yong)前(qian)(qian)(qian)加(jia)入 2mL 雙(shuang)(shuang)蒸(zheng)水(shui)充分(fen)(fen)溶(rong)解(jie)備用(yong)(yong)(yong)(yong)；用(yong)(yong)(yong)(yong)不(bu)完的(de)試(shi)(shi)(shi)(shi)(shi)劑(ji)仍(reng) 4℃保(bao)(bao)(bao)存(cun)(cun)；試(shi)(shi)(shi)(shi)(shi)劑(ji)四：粉(fen)劑(ji)×1 瓶(ping)(ping)(ping)，4℃保(bao)(bao)(bao)存(cun)(cun)；臨(lin)(lin)(lin)用(yong)(yong)(yong)(yong)每(mei)瓶(ping)(ping)(ping)前(qian)(qian)(qian)加(jia)入 3mL 雙(shuang)(shuang)蒸(zheng)水(shui)充分(fen)(fen)溶(rong)解(jie)備用(yong)(yong)(yong)(yong)；用(yong)(yong)(yong)(yong)不(bu)完的(de)試(shi)(shi)(shi)(shi)(shi)劑(ji)仍(reng) 4℃保(bao)(bao)(bao)存(cun)(cun)；試(shi)(shi)(shi)(shi)(shi)劑(ji)五：粉(fen)劑(ji)×1 瓶(ping)(ping)(ping)，-20℃保(bao)(bao)(bao)存(cun)(cun)；臨(lin)(lin)(lin)用(yong)(yong)(yong)(yong)前(qian)(qian)(qian)每(mei)瓶(ping)(ping)(ping)加(jia)入 3mL 雙(shuang)(shuang)蒸(zheng)水(shui)充分(fen)(fen)溶(rong)解(jie)備用(yong)(yong)(yong)(yong)；用(yong)(yong)(yong)(yong)不(bu)完的(de)試(shi)(shi)(shi)(shi)(shi)劑(ji)仍(reng)-20℃保(bao)(bao)(bao)存(cun)(cun)；試(shi)(shi)(shi)(shi)(shi)劑(ji)六：粉(fen)劑(ji)×1 支(zhi)(zhi)，-20℃保(bao)(bao)(bao)存(cun)(cun)；臨(lin)(lin)(lin)用(yong)(yong)(yong)(yong)前(qian)(qian)(qian)每(mei)支(zhi)(zhi)加(jia)入 250μL 雙(shuang)(shuang)蒸(zheng)水(shui)充分(fen)(fen)溶(rong)解(jie)備用(yong)(yong)(yong)(yong)；用(yong)(yong)(yong)(yong)不(bu)完的(de)試(shi)(shi)(shi)(shi)(shi)劑(ji) 4℃保(bao)(bao)(bao)存(cun)(cun)；試(shi)(shi)(shi)(shi)(shi)劑(ji)七：粉(fen)劑(ji)×1 支(zhi)(zhi)，-20℃保(bao)(bao)(bao)存(cun)(cun)；臨(lin)(lin)(lin)用(yong)(yong)(yong)(yong)前(qian)(qian)(qian)每(mei)支(zhi)(zhi)加(jia)入 250μL 雙(shuang)(shuang)蒸(zheng)水(shui)充分(fen)(fen)溶(rong)解(jie)備用(yong)(yong)(yong)(yong)；用(yong)(yong)(yong)(yong)不(bu)完的(de)試(shi)(shi)(shi)(shi)(shi)劑(ji) 4℃保(bao)(bao)(bao)存(cun)(cun)；