產品名稱:植物中(zhong)脂氧合(he)酶 (LOX)活性(xing)檢測試劑盒
產品型號:BC0325
產品特點:植物中脂氧合酶 (LOX)活性檢測試劑盒測定意義:LOX廣泛存在于植物組織中,催化不飽和脂肪酸氧化反應,導致膜脂過氧化。在植物的生長發育、成熟衰老及逆境脅迫過程中具有重要作用。測定原理:LOX催化亞油酸氧化,氧化產物在234nm處有特征吸收峰;測定234nm吸光度增加速率,來計算LOX活性。
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植物中脂氧合酶 (LOX)活性檢測試劑盒
LOX 廣泛存在于植物(wu)組織中(zhong),催(cui)化(hua)不飽和(he)脂肪酸(suan)氧化(hua)反(fan)應,導致膜脂過(guo)氧化(hua)。在植物(wu)的生長發育、成熟衰(shuai)老及逆境脅(xie)迫過(guo)程中(zhong)具有(you)重要作用(yong)。 LOX 催(cui)化(hua)亞(ya)油酸(suan)氧化(hua),氧化(hua)產物(wu)在 234nm 處有(you)特(te)征吸收峰;測定 234nm 吸光度增加(jia)速率,來計算 LOX 活性。
植物中脂氧合酶 (LOX)活性檢測試劑盒
一、粗酶(mei)液(ye)(ye)提取:按照(zhao)組織質量(liang)(liang)(liang)(g):試(shi)(shi)劑(ji)(ji)一體(ti)(ti)(ti)積(ji)(ji)(mL)為(wei)(wei) 1:5~10 的比(bi)(bi)(bi)例(建議稱取約 0.1g 組織,加(jia)(jia)入(ru) 1mL 試(shi)(shi)劑(ji)(ji)一)進行(xing)冰浴(yu)勻漿。16000g,4℃離(li)心 20min,取上(shang)(shang)清(qing)(qing)置(zhi)冰上(shang)(shang)待測(ce)(ce)(ce)(ce)。二(er)、測(ce)(ce)(ce)(ce)定(ding)(ding): 1. 分(fen)光(guang)光(guang)度計(ji)/酶(mei)標儀(yi)預(yu)熱(re) 30 min 以(yi)上(shang)(shang),調節波(bo)長到 234 nm,蒸餾水調零。 2. 試(shi)(shi)劑(ji)(ji)二(er)在(zai)(zai) 25℃水浴(yu)中(zhong)(zhong)預(yu)熱(re) 30 min 以(yi)上(shang)(shang)。 3. 空白(bai)(bai)管(guan):依次(ci)在(zai)(zai)微(wei)(wei)量(liang)(liang)(liang)石英(ying)比(bi)(bi)(bi)色(se)(se)皿/96 孔板(ban)中(zhong)(zhong)加(jia)(jia)入(ru) 20μL 蒸餾水、160μL 試(shi)(shi)劑(ji)(ji)二(er)和(he)(he) 20μL 試(shi)(shi)劑(ji)(ji)三(san),迅(xun)速混勻后(hou)(hou)于 234nm 比(bi)(bi)(bi)色(se)(se),記(ji)錄 15s 和(he)(he) 75s 的吸光(guang)值(zhi),分(fen)別記(ji)為(wei)(wei) A1 和(he)(he) A2。 4. 測(ce)(ce)(ce)(ce)定(ding)(ding)管(guan):依次(ci)在(zai)(zai)微(wei)(wei)量(liang)(liang)(liang)石英(ying)比(bi)(bi)(bi)色(se)(se)皿/96 孔板(ban)中(zhong)(zhong)加(jia)(jia)入(ru) 20μL 上(shang)(shang)清(qing)(qing)液(ye)(ye)、160μL 試(shi)(shi)劑(ji)(ji)二(er)和(he)(he) 20μL 試(shi)(shi)劑(ji)(ji)三(san),迅(xun)速混勻后(hou)(hou)于 234nm 比(bi)(bi)(bi)色(se)(se),記(ji)錄 15s 和(he)(he) 75s 的吸光(guang)值(zhi),分(fen)別記(ji)為(wei)(wei) A3 和(he)(he) A4。三(san)、LOX 活性計(ji)算(suan): a. 使用(yong)(yong)微(wei)(wei)量(liang)(liang)(liang)石英(ying)比(bi)(bi)(bi)色(se)(se)皿測(ce)(ce)(ce)(ce)定(ding)(ding)的計(ji)算(suan)公式(shi)如下(1) 按照(zhao)蛋(dan)(dan)(dan)白(bai)(bai)濃(nong)度計(ji)算(suan)活性單(dan)位(wei)(wei)定(ding)(ding)義(yi):25℃中(zhong)(zhong)每(mei)(mei)毫克(ke)(ke)蛋(dan)(dan)(dan)白(bai)(bai)每(mei)(mei)分(fen)鐘(zhong)催化(hua)吸光(guang)值(zhi)變化(hua) 0.001 個(ge)單(dan)位(wei)(wei)為(wei)(wei) 1 個(ge)酶(mei)活單(dan)位(wei)(wei)。 LOX (U/mg prot) = [(A4-A3)-(A2-A1)]×V 反(fan)總(zong)(zong)(zong)÷(Cpr×V 樣(yang)(yang))÷T×1000 = 10000×[(A4-A3)-(A2-A1)]÷Cpr Solarbio Beijing Solarbio Science & Technology Co., Ltd /82 Http://www.solarbio。。com 第 2頁,共(gong) 2 頁(2) 按照(zhao)樣(yang)(yang)本(ben)質量(liang)(liang)(liang)計(ji)算(suan)活性單(dan)位(wei)(wei)定(ding)(ding)義(yi):25℃中(zhong)(zhong)每(mei)(mei)克(ke)(ke)組織每(mei)(mei)分(fen)鐘(zhong)催化(hua)吸光(guang)值(zhi)變化(hua) 0.001 個(ge)單(dan)位(wei)(wei)為(wei)(wei) 1 個(ge)酶(mei)活單(dan)位(wei)(wei)。 LOX (U/g) = [(A4-A3)-(A2-A1)]×V 反(fan)總(zong)(zong)(zong)÷(W×V 樣(yang)(yang)÷V 樣(yang)(yang)總(zong)(zong)(zong))÷T×1000 = 10000×[(A4-A3)-(A2-A1)]÷W Cpr:上(shang)(shang)清(qing)(qing)液(ye)(ye)蛋(dan)(dan)(dan)白(bai)(bai)濃(nong)度,mg/mL,需(xu)另外測(ce)(ce)(ce)(ce)定(ding)(ding),建議使用(yong)(yong)本(ben)公司 BCA 蛋(dan)(dan)(dan)白(bai)(bai)質含量(liang)(liang)(liang)測(ce)(ce)(ce)(ce)定(ding)(ding)試(shi)(shi)劑(ji)(ji)盒; V 反(fan)總(zong)(zong)(zong):反(fan)應(ying)(ying)體(ti)(ti)(ti)系(xi)總(zong)(zong)(zong)體(ti)(ti)(ti)積(ji)(ji),200μL=2×10-4L;V 樣(yang)(yang):加(jia)(jia)入(ru)反(fan)應(ying)(ying)體(ti)(ti)(ti)系(xi)中(zhong)(zhong)上(shang)(shang)清(qing)(qing)液(ye)(ye)體(ti)(ti)(ti)積(ji)(ji),20μL=0.02mL;W :樣(yang)(yang)品質量(liang)(liang)(liang),g;V 樣(yang)(yang)總(zong)(zong)(zong):上(shang)(shang)清(qing)(qing)液(ye)(ye)總(zong)(zong)(zong)體(ti)(ti)(ti)積(ji)(ji),1mL;T:反(fan)應(ying)(ying)時間。 b. 使用(yong)(yong) 96 孔板(ban)測(ce)(ce)(ce)(ce)定(ding)(ding)的計(ji)算(suan)公式(shi)如下(1) 按照(zhao)蛋(dan)(dan)(dan)白(bai)(bai)濃(nong)度計(ji)算(suan) 活性單(dan)位(wei)(wei)定(ding)(ding)義(yi):25℃中(zhong)(zhong)每(mei)(mei)毫克(ke)(ke)蛋(dan)(dan)(dan)白(bai)(bai)每(mei)(mei)分(fen)鐘(zhong)催化(hua)吸光(guang)值(zhi)變化(hua) 0.001 個(ge)單(dan)位(wei)(wei)為(wei)(wei) 1 個(ge)酶(mei)活單(dan)位(wei)(wei)。 LOX (U/mg prot) = [(A4-A3)-(A2-A1)]×V 反(fan)總(zong)(zong)(zong)÷(Cpr×V 樣(yang)(yang))÷T×1000 = 10 000×[(A4-A3)-(A2-A1)]÷Cpr (2) 按照(zhao)樣(yang)(yang)本(ben)質量(liang)(liang)(liang)計(ji)算(suan) 活性單(dan)位(wei)(wei)定(ding)(ding)義(yi):25℃中(zhong)(zhong)每(mei)(mei)克(ke)(ke)組織每(mei)(mei)分(fen)鐘(zhong)催化(hua)吸光(guang)值(zhi)變化(hua) 0.001 個(ge)單(dan)位(wei)(wei)為(wei)(wei) 1 個(ge)酶(mei)活單(dan)位(wei)(wei)。 LOX (U/g) = [(A4-A3)-(A2-A1)]×V 反(fan)總(zong)(zong)(zong)÷(W×V 樣(yang)(yang)÷V 樣(yang)(yang)總(zong)(zong)(zong))÷T×1000 = 10 000×[(A4-A3)-(A2-A1)]÷W Cpr:上(shang)(shang)清(qing)(qing)液(ye)(ye)蛋(dan)(dan)(dan)白(bai)(bai)濃(nong)度,mg/mL,需(xu)另外測(ce)(ce)(ce)(ce)定(ding)(ding),建議使用(yong)(yong)本(ben)公司 BCA 蛋(dan)(dan)(dan)白(bai)(bai)質含量(liang)(liang)(liang)測(ce)(ce)(ce)(ce)定(ding)(ding)試(shi)(shi)劑(ji)(ji)盒; V 反(fan)總(zong)(zong)(zong):反(fan)應(ying)(ying)體(ti)(ti)(ti)系(xi)總(zong)(zong)(zong)體(ti)(ti)(ti)積(ji)(ji),200μL=2×10 -4L;V 樣(yang)(yang):加(jia)(jia)入(ru)反(fan)應(ying)(ying)體(ti)(ti)(ti)系(xi)中(zhong)(zhong)上(shang)(shang)清(qing)(qing)液(ye)(ye)體(ti)(ti)(ti)積(ji)(ji),20μL=0.02mL;W :樣(yang)(yang)品質量(liang)(liang)(liang),g;V 樣(yang)(yang)總(zong)(zong)(zong):上(shang)(shang)清(qing)(qing)液(ye)(ye)總(zong)(zong)(zong)體(ti)(ti)(ti)積(ji)(ji),1mL;T:反(fan)應(ying)(ying)時間。
植物中脂氧合酶 (LOX)活性檢測試劑盒
試劑(ji)(ji)一(yi):液(ye)體×1 瓶(ping)(ping),4℃保(bao)存。試劑(ji)(ji)二(er):液(ye)體×1 瓶(ping)(ping),4℃保(bao)存。試劑(ji)(ji)三:粉劑(ji)(ji)×1 瓶(ping)(ping),4℃保(bao)存。臨用前加入 2mL 試劑(ji)(ji)二(er),充分溶解。
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