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MTT一般現用現配,過濾后4℃,避光保存2周有效,或保存在-20℃長期保存;避免反復凍融,最好小劑量分裝,用避光袋、錫箔紙包裹;一般把MTT粉分裝在EP管,再現用現配,當MTT變?yōu)榛揖G色時絕不能再用; MTT致癌,用時最好帶透明薄膜手套。
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培養(yǎng)基保存于4℃冰箱中,培養(yǎng)基內CO2 會逐漸溢出,造成培養(yǎng)基越來越偏堿性。而培養(yǎng)基中酸堿指示劑(通常為phenol red)的顏色也會隨堿性增加而更偏暗紅。
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培養(yǎng)基偏堿后再用于細胞培養(yǎng)將造成細胞生長停滯或死亡。培養(yǎng)基偏堿時,可以通入無菌過濾的CO2,以調整pH值。
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根據細胞類型、培養(yǎng)方式和生產工藝等特點所定制的培養(yǎng)基,即個性化培養(yǎng)基。個性化培養(yǎng)基在國外生物制藥企業(yè)被普遍采用,個性化培養(yǎng)基可以為細胞生長提供充足的營養(yǎng)物質,能提高細胞的生長速率、培養(yǎng)密度、以及延長細胞維持時間;
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同時對貼壁細胞而言,能增加細胞的貼壁性,并降低培養(yǎng)過程中剪切力對細胞的損傷;個性化培養(yǎng)基可以根據各戶的特殊需求減少或不使用動物源成分,從而使生物制品安全性更有保障。
全氟消解罐又叫COD樣品消解儀、消解器、高壓消解罐、在海洋研究所用于海洋樣品檢測尤為廣泛。應用于氣象,液相,等離子光質譜儀,原子吸收和原子熒光等化學分析方法的樣品前處理,用于消解農殘,食品,稀土,水產品,有機物種的Pb,Cu,Zn,Ga,Rb,Hg等重金屬。案例:清華大學:COD檢測。
1.耐高低溫性:可使用溫度-200℃~+250℃。建議在烘箱中180度內使用(全氟消解罐)
2.使用方便精密設備加工內壁光滑,不掛水,不混配,密封性能好
3.消解效率高,能力強,可消解許多傳統方法難以消解的樣品
4.消耗酸溶劑少,空白值低,提高分析的準確度和精密度
5.采用全聚四氟乙烯材質加工,避免了在實驗中,混入重金屬,從而污染樣品,防污染:金屬元素空白值低。
6.外觀純白色。
7.耐腐蝕:耐強酸、強堿、王水和各種有機溶劑,且無溶出、吸附和析出現象。
8.絕緣性:不受環(huán)境及頻率的影響,介質損耗小,擊穿電壓高。
9.耐大氣老化,耐輻照和較低的滲透性。
10.自潤滑性:具有塑料中最小的摩擦系數。
11.表面不粘性:是一種表面能最小的固體材料。
12.機械性質較軟,具有非常低的表面能。
13.無毒害:具有生理惰性
銷售部:
聯系人: 季小姐
TEL:138 138 88374 025-8559 7772
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【Discovery 專業(yè)型分析天平 DV 分析天平(內校)0.00001/0.0001g的參數說明】 | |||||||||||||||||||||||
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人棘球蚴IgM抗體酶聯免疫分析(ELISA)
試劑盒使用說明書
本試劑僅供研究使用 目的:本試劑盒用于測定人血清,血漿及相關液體樣本中棘球蚴IgM抗體的含量。
實驗原理:
本試劑盒應用雙抗原夾心法測定標本中人棘球蚴IgM抗體水平。用純化的人棘球蚴IgM抗原包被微孔板,制成固相抗原,往包被抗原的微孔中依次加入棘球蚴IgM抗體,再與HRP標記的棘球蚴IgM抗原結合,形成抗原-抗體-酶標抗原復合物,經過洗滌后加底物TMB顯色。TMB在HRP酶的催化下轉化成藍色,并在酸的作用下轉化成最終的黃色。顏色的深淺和樣品中的棘球蚴IgM抗體呈正相關。用酶標儀在450nm波長下測定吸光度(OD值),通過標準曲線計算樣品中人棘球蚴IgM抗體濃度。
試劑盒組成:
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樣本處理及要求:
1. 血清:室溫血液自然凝固10-20分鐘,離心20分鐘左右(2000-3000轉/分)。仔細收集上清,保存過程中如出現沉淀,應再次離心。
2. 血漿:應根據標本的要求選擇EDTA或檸檬酸鈉作為抗凝劑,混合10-20分鐘后,離心20分鐘左右(2000-3000轉/分)。仔細收集上清,保存過程中如有沉淀形成,應該再次離心。
3. 尿液:用無菌管收集,離心20分鐘左右(2000-3000轉/分)。仔細收集上清,保存過程中如有沉淀形成,應再次離心。胸腹水、腦脊液參照實行。
4. 細胞培養(yǎng)上清:檢測分泌性的成份時,用無菌管收集。離心20分鐘左右(2000-3000轉/分)。仔細收集上清。檢測細胞內的成份時,用PBS(PH7.2-7.4)稀釋細胞懸液,細胞濃度達到100萬/ml左右。通過反復凍融,以使細胞破壞并放出細胞內成份。離心20分鐘左右(2000-3000轉/分)。仔細收集上清。保存過程中如有沉淀形成,應再次離心。
5. 組織標本:切割標本后,稱取重量。加入一定量的PBS,PH7.4。用液氮迅速冷凍保存?zhèn)溆。標本融化后仍然保?/span>2-8℃的溫度。加入一定量的PBS(PH7.4),用手工或勻漿器將標本勻漿充分。離心20分鐘左右(2000-3000轉/分)。仔細收集上清。分裝后一份待檢測,其余冷凍備用。
6. 標本采集后盡早進行提取,提取按相關文獻進行,提取后應盡快進行實驗。若不能馬上進行試驗,可將標本放于-20℃保存,但應避免反復凍融.
7. 不能檢測含NaN3的樣品,因NaN3抑制辣根過氧化物酶的(HRP)活性。
操作步驟
1. 標準品的稀釋與加樣:在酶標包被板上設標準品孔10孔,在、第二孔中分別加標準品100μl,然后在、第二孔中加標準品稀釋液50μl,混勻;然后從孔、第二孔中各取100μl分別加到第三孔和第四孔,再在第三、第四孔分別加標準品稀釋液50μl,混勻;然后在第三孔和第四孔中先各取50μl棄掉,再各取50μl分別加到第五、第六孔中,再在第五、第六孔中分別加標準品稀釋液50ul,混勻;混勻后從第五、第六孔中各取50μl分別加到第七、第八孔中,再在第七、第八孔中分別加標準品稀釋液50μl,混勻后從第七、第八孔中分別取50μl加到第九、第十孔中,再在第九第十孔分別加標準品稀釋液50μl,混勻后從第九第十孔中各取50μl棄掉。(稀釋后各孔加樣量都為50μl,濃度分別為90pg/ml,60pg/ml ,30 pg/ml,15pg/ml , 7.5pg/ml。
2. 加樣:分別設空白孔(空白對照孔不加樣品及酶標試劑,其余各步操作相同)、待測樣品孔。在酶標包被板上待測樣品孔中先加樣品稀釋液40μl,然后再加待測樣品10μl(樣品最終稀釋度為5倍)。加樣將樣品加于酶標板孔底部,盡量不觸及孔壁,輕輕晃動混勻。
3. 溫育:用封板膜封板后置37℃溫育30分鐘。
4. 配液:將30(48T的20倍)倍濃縮洗滌液用蒸餾水30(48T的20倍)倍稀釋后備用。
5. 洗滌:小心揭掉封板膜,棄去液體,甩干,每孔加滿洗滌液,靜置30秒后棄去,如此重復5次,拍干。
6. 加酶:每孔加入酶標試劑50μl,空白孔除外。
7. 溫育:操作同3。
8. 洗滌:操作同5。
9. 顯色:每孔先加入顯色劑A50μl,再加入顯色劑B50μl,輕輕震蕩混勻,37℃避光顯色15分鐘.
10. 終止:每孔加終止液50μl,終止反應(此時藍色立轉黃色)。
11. 測定:以空白空調零,450nm波長依序測量各孔的吸光度(OD值)。 測定應在加終止液后15分鐘以內進行。
注意事項:
1. 試劑盒從冷藏環(huán)境中取出應在室溫平衡15-30分鐘后方可使用,酶標包被板開封后如未用完,板條應裝入密封袋中保存。
2. 濃洗滌液可能會有結晶析出,稀釋時可在水浴中加溫助溶,洗滌時不影響結果。
3. 各步加樣均應使用加樣器,并經常校對其性,以避免試驗誤差。一次加樣時間控制在5分鐘內,如標本數量多,推薦使用排槍加樣。
4. 請每次測定的同時做標準曲線,做復孔。如標本中待測物質含量過高(樣本OD值大于標準品孔孔的OD值),請先用樣品稀釋液稀釋一定倍數(n倍)后再測定,計算時請乘以總稀釋倍數(×n×5)。
5. 封板膜只限一次性使用,以避免交叉污染。
6. 底物請避光保存。
7. 嚴格按照說明書的操作進行,試驗結果判定必須以酶標儀讀數為準.
8. 所有樣品,洗滌液和各種廢棄物都應按傳染物處理。
9. 本試劑不同批號組分不得混用。
10. 如與英文說明書有異,以英文說明書為準。
計算:
以標準物的濃度為橫坐標,OD值為縱坐標,
在坐標紙上繪出標準曲線,根據樣品的OD
值由標準曲線查出相應的濃度;再乘以稀釋
倍數;或用標準物的濃度與OD值計算出標
準曲線的直線回歸方程式,將樣品的OD值
代入方程式,計算出樣品濃度,再乘以稀釋
倍數,即為樣品的實際濃度。
(此圖僅供參考)
試劑盒性能:
1.樣品線性回歸與預期濃度相關系數R值為0.92以上。
2.批內與批見應分別小于9%和15%
檢測范圍:
3pg/ml -100 pg/ml
保存條件及期:
1.試劑盒保存:;2-8℃。
2.期:6個月
FOR RESEARCH USE ONLY
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Drug Names
Generic Name:Human echinococcosis IgM antibody (ECH-IgM) ELISA Kit.
Purpose
This kit allows for the determination of ECH-IgM concentrations in Human serum, blood plasma, and other biological fluids.
Principle of the assay
The kit assay Human ECH-IgM level in the sample,use Purified Human ECH-IgM antigen to coat microtiter plate wells, make solid-phase antigen, then add ECH-IgM to wells, Combined ECH-IgM antigen which With HRP labeled , become antigen - antibody - enzyme- antigen complex, after washing Completely, Add TMB substrate solution,TMB substrate becomes blue color At HRP enzyme-catalyzed, reaction is terminated by the addition of a sulphuric acid solution and the color change is measured spectrophotometrically at a wavelength of 450 nm. The concentration of ECH-IgM in the samples is then determined by comparing the O.D. of the samples to the standard curve.
Materials provided with the kit
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| (20ml×20 fold) ×1bottle | (20ml×30 fold) | |
Specimen requirements
1. serum- coagulation at room temperature 10-20 mins,centrifugation 20-min at the speed of 2000-3000 r.p.m. remove supernatant, If precipitation appeared, Centrifugal again.
2. plasma-use suited EDTA or citrate plasma as an anticoagulant,mix 10-20 mins ,centrifugation 20-min at the speed of 2000-3000 r.p.m. remove supernatant, If precipitation appeared, Centrifugal again.
3. Urine-collect sue a sterile container, centrifugation 20-min at the speed of 2000-3000 r.p.m. remove supernatant, If precipitation appeared, Centrifugal again. The Operation of Hydrothorax and cerebrospinal fluid Reference to it.
4. cell culture supernatant-detect secretory components, collect sue a sterile container, centrifugation 20-min at the speed of 2000-3000 r.p.m. remove supernatant,detect the composition of cells, Dilut cell suspension with PBS(PH7.2-7.4), Cell concentration reached 1 million / ml, repeated freeze-thaw cycles, damage cells and release of intracellular components, centrifugation 20-min at the speed of 2000-3000 r.p.m. remove supernatant, If precipitation appeared, Centrifugal again.
5. Tissue samples- After cutting samples, check the weight,add PBS(PH7.2-7.4), Rapidly frozen with liquid nitrogen, maintain samples at 2-8℃ after melting,add PBS(PH7.4), Homogenized by hand or Grinders, centrifugation 20-min at the speed of 2000-3000 r.p.m. remove supernatant.
6. extract as soon as possible after Specimen collection,and according to the relevant literature, and should be experiment as soon as possible after the extraction. If it can’t, specimen can be kept in -20 ℃ to preserve, Avoid repeated freeze-thaw cycles.
7. Can’t detect the sample which contain NaN3, because NaN3 inhibits HRP active.
Assay procedure
1.Dilute and add sample to Standard: set 10 Standard wells on the ELISA plates coated, add Standard 100μl to the first and the second well, then add Standard dilution 50μl to the first and the second well, mix; take out 100μl form the first and the second well then add it to the third and the forth well separately. then add Standard dilution 50μl to the third and the forth well ,mix ; then take out 50μl from the third and the forth well discard, add 50μl to the fifth and the sixth well ,then add Standard dilution 50μl to the fifth and the sixth well, mix ; take out 50μl from the fifth and the sixth well and add to the seventh and the eighth well, then add Standard dilution 50μl to the seventh and the eighth well ,mix ; take out 50μl from the seventh and the eighth well and add to the ninth and the tenth well, add Standard dilution 50μl to the ninth and the tenth well, mix , take out 50μl from the ninth and the tenth well discard(add Sample 50μl to each well after Diluting ,(density: 90pg/ml,60pg/ml ,30 pg/ml,15pg/ml , 7.5pg/ml)
2.add sample:Set blank wells separately (blank comparison wells don’t add sample and HRP-Conjugate reagent, other each step operation is same). testing sample well. add Sample dilution 40μl to testing sample well, then add testing sample 10μl (sample final dilution is 5-fold), add sample to wells , don’t touch the well wall as far as possible, and Gently mix.
3.Incubate: After closing plate with Closure plate membrane ,incubate for 30 min at 37℃.
4.Configurate liquid: 30-fold (or 20-fold)wash solution diluted 30-fold (or 20-fold) with distilled water and reserve.
5.washing:Uncover Closure plate membrane, discard Liquid, dry by swing, add washing buffer to every well, still for 30s then drain, repeat 5 times, dry by pat.
6.add enzyme:Add HRP-Conjugate reagent 50μl to each well, except blank well.
7.incubate:Operation with 3.
8.washing:Operation with 5.
9.color:Add Chromogen Solution A 50ul and Chromogen Solution B to each well, evade the light preservation for 15 min at 37℃
10.Stop the reaction:Add Stop Solution50μl to each well, Stop the reaction(the blue color change to yellow color).
11.assay:take blank well as zero , Read absorbance at 450nm after Adding Stop Solution and within 15min.
Important notes
1. The kit takes out from the refrigeration environment should be balanced 15-30 minutes in the room temperature, ELISA plates coated if has not use up after opened, the plate should be stored in Sealed bag.
2. washing buffer will Crystallization separation, it can be heated the water helps dissolve when dilute . Washing does not affect the result.
3. add Sample with sampler Each step, And proofread its accuracy frequently, avoids the experimental error. add sample within 5 mins, if the number of sample is much , recommend to use Volley .
4. if the testing material content is excessively higher (The sample OD is bigger than the first standard well ),please dilute Sample (n-fold), Please diluente and multiplied by the dilution factor.(×n×5).
5. Closure plate membrane only limits the disposable use, to avoid cross-contamination.
6. The substrate evade the light preservation.
7. Please according to use instruction strictly, The test result determination must take the microtiter plate reader as a standard.
8. All samples, washing buffer and each kind of reject should according to infective material process.
9. Do not mix reagents with those from other lots.
Take the standard density as the horizontal, the OD value for the vertical ,draw the standard curve on graph paper, Find out the corresponding density according to the sample OD value by the Sample curve, multiplied by the dilution multiple, or calculate the straight line regression equation of the standard curve with the standard density and the OD value ,with the sample OD value in the equation, calculate the sample density, multiplied by the dilution factor, the result is the sample actual density.
Calculate
This chart for reference only
Assay range
3pg/ml -100 pg/ml
Storage and validity
1.Storage: 2-8℃.
2.validity: six months.
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技術參數★ 溫控系統:165℃±1.0℃★ 消解時間:10分鐘★ 測定范圍:50-800mg/L★ 測定時間: 20分鐘同時測定9個水樣★ 抗氯干擾:[CL-]<1000 mg/L★ 度: COD=300 mg/L 相對誤差V≤10 %★ 外型尺寸:210×160×67mm★ 內存1條標準曲線,可自行修定,具有掉電保留功能★ 冷光源、窄帶干涉、光源壽命長★ 定時系統:三個定時開關,自動提醒其他說明標準配置:反應器(消解器)、冷卻槽架、專用固體試劑、反應管數支。
(1) 可以設定消解時間,消解完畢后,儀器自動停止加熱,可無人看管
(1) 可以設定消解時間,消解完畢后,儀器自動停止加熱,可無人看管
(2) 樣品消解完畢后,儀器風機繼續(xù)工作半小時,輔助樣品冷卻。
(3) 節(jié)約用電、用水,提高了效率,增強了儀器的安全性。