減速機(jī)是一種動(dòng)力傳達(dá)機(jī)構(gòu)，利用齒輪的速度轉(zhuǎn)換器，將電機(jī)（馬達(dá)）的回轉(zhuǎn)數(shù)減速到所要的回轉(zhuǎn)數(shù)，并得到較大轉(zhuǎn)矩的機(jī)構(gòu)。在目前用于傳遞動(dòng)力與運(yùn)動(dòng)的機(jī)構(gòu)中，減速機(jī)的應(yīng)用范圍相當(dāng)廣泛。幾乎在各式機(jī)械的傳動(dòng)系統(tǒng)中都可以見到它的蹤跡，從交通工具的船舶、汽車、機(jī)車，建筑用的重型機(jī)具，機(jī)械工業(yè)所用的加工機(jī)具及自動(dòng)化生產(chǎn)設(shè)備，到日常生活中常見的家電，鐘表等等.其應(yīng)用從大動(dòng)力的傳輸工作，到小負(fù)荷，精確的角度傳輸都可以見到減速機(jī)的應(yīng)用，且在工業(yè)應(yīng)用上，減速機(jī)具有減速及增加轉(zhuǎn)矩功能。因此廣泛應(yīng)用在速度與扭矩的轉(zhuǎn)換設(shè)備。減速機(jī)的作用主要有： 1）降速同時(shí)提高輸出扭矩，扭矩輸出比例按電機(jī)輸出乘減速比，但要注意不能超出減速機(jī)額定扭矩。 2）減速同時(shí)降低了負(fù)載的慣量，慣量的減少為減速比的平方。大家可以看一下一般電機(jī)都有一個(gè)慣量數(shù)值。 減速機(jī)的工作原理 減速機(jī)一般用于低轉(zhuǎn)速大扭矩的傳動(dòng)設(shè)備，把電動(dòng)機(jī).內(nèi)燃機(jī)或其它高速運(yùn)轉(zhuǎn)的動(dòng)力通過減速機(jī)的輸入軸上的齒數(shù)少的齒輪嚙合輸出軸上的大齒輪來達(dá)到減速的目的，普通的減速機(jī)也會(huì)有幾對(duì)相同原理齒輪達(dá)到理想的減速效果，大小齒輪的齒數(shù)之比，就是傳動(dòng)比。[編輯本段]減速機(jī)的種類 減速機(jī)是一種相對(duì)精密的機(jī)械，使用它的目的是降低轉(zhuǎn)速，增加轉(zhuǎn)矩。它的種類繁多，型號(hào)各異，不同種類有不同的用途。減速器的種類繁多，按照傳動(dòng)類型可分為齒輪減速器、蝸桿減速器和行星齒輪減速器；按照傳動(dòng)級(jí)數(shù)不同可分為單級(jí)和多級(jí)減速器；按照齒輪形狀可分為圓柱齒輪減速器、圓錐齒輪減速器和圓錐－圓柱齒輪減速器；按照傳動(dòng)的布置形式又可分為展開式、分流式和同軸式減速器。以下是常用的減速機(jī)分類： ⑴擺線針輪減速機(jī) ⑵硬齒面圓柱齒輪減速器 ⑶行星齒輪減速機(jī) ⑷軟齒面減速機(jī) ⑸三環(huán)減速機(jī) ⑹起重機(jī)減速機(jī) ⑺蝸桿減速機(jī) ⑻軸裝式硬齒面減速機(jī) ⑼無級(jí)變速器 蝸輪蝸桿減速機(jī)的主要特點(diǎn)是具有反向自鎖功能，可以有較大的減速比，輸入軸和輸出軸不在同一軸線上，也不在同一平面上。但是一般體積較大，傳動(dòng)效率不高，精度不高。諧波減速機(jī)的諧波傳動(dòng)是利用柔性元件可控的彈性變形來傳遞運(yùn)動(dòng)和動(dòng)力的，體積不大、精度很高，但缺點(diǎn)是柔輪壽命有限、不耐沖擊，剛性與金屬件相比較差。輸入轉(zhuǎn)速不能太高。行星減速機(jī)其優(yōu)點(diǎn)是結(jié)構(gòu)比較緊湊，回程間隙小、精度較高，使用壽命很長(zhǎng)，額定輸出扭矩可以做的很大。但價(jià)格略貴。 擺線減速機(jī)特點(diǎn) 行星擺線減速機(jī)是一種應(yīng)用行星傳動(dòng)原理，采用擺線針輪嚙合，設(shè)計(jì)、結(jié)構(gòu)新穎。這種減速機(jī)在絕大多數(shù)情況下已替代兩級(jí)、三級(jí)普通圓柱齒輪減速機(jī)及圓柱蝸桿減速機(jī)，在軍工、航天、冶金、礦、石油、化工、船舶、輕工、食品、紡織、印染、制藥、橡膠、塑料、及起重運(yùn)輸?shù)确矫娴玫饺找鎻V泛的應(yīng)用。 一、產(chǎn)品特點(diǎn) 1.傳動(dòng)比大。一級(jí)減速時(shí)傳動(dòng)比為1/6－－1/87。兩級(jí)減速時(shí)傳動(dòng)比為1/99－－1/7569；三級(jí)傳動(dòng)時(shí)傳動(dòng)比為1/5841－－1/658503。另外根據(jù)需要還可以采用多級(jí)組合，速比達(dá)到指定大。 2.傳動(dòng)效率高。由于嚙合部位采用了滾動(dòng)嚙合，一般一級(jí)傳動(dòng)效率為90%－－95%。 3.結(jié)構(gòu)緊湊，體積小，重量輕。體積和普通圓柱齒輪減速機(jī)相比可減小2/1－－2/3。 4.故障少，壽命長(zhǎng)。主要傳動(dòng)嚙合件使用軸承鋼磨削制造，因此機(jī)械性能與耐磨性能均佳，又因其為滾動(dòng)摩擦，因而故障少，壽命長(zhǎng)。 5.運(yùn)轉(zhuǎn)平穩(wěn)。因傳動(dòng)過程中為多齒嚙合，所以使之運(yùn)轉(zhuǎn)平穩(wěn)，噪聲低。 6.拆裝方便，容易維修。 7.過載能力強(qiáng)，耐沖擊，慣性力矩小，適用于起動(dòng)頻繁和正反轉(zhuǎn)運(yùn)轉(zhuǎn)的特點(diǎn)。 二、技術(shù)規(guī)格 1、機(jī)型號(hào): 按傳動(dòng)比分為:一級(jí)、二級(jí)、三級(jí)。 一級(jí)有十三種機(jī)型：0，1，2，3，4，5，6，7，8，9，10，11，12。 兩級(jí)有14種機(jī)型：00，20；32，42，53，63，64，74，84，85，95，106，117，128。 三級(jí)有8種機(jī)型：420，742，842，853，953，1063，1174，1285。 按結(jié)構(gòu)型式分為：臥式、立式、雙軸型、直聯(lián)型四種。 2、傳動(dòng)比： 一級(jí)減速的傳動(dòng)比有：9，11，17，21，23，25，29，35，43，47，59，71，87。 兩級(jí)減速的傳動(dòng)比有：99，121，187，289，319，385，473，493，595，649，731，841，1003，1225， 1505，1849，2065，2537，3045，3481，5133。 三級(jí)減速的傳動(dòng)比有：5841-658530[編輯本段]減速機(jī)的發(fā)展 20世紀(jì)70－80年代，世界上減速器技術(shù)有了很大的發(fā)展，且與新技術(shù)革命的發(fā)展緊密結(jié)合。通用減速器的發(fā)展趨勢(shì)如下： ①高水平、高性能。圓柱齒輪普遍采用滲碳淬火、磨齒，承載能力提高4倍以上，體積小、重量輕、噪聲低、效率高、性高。 ②積木式組合設(shè)計(jì)；緟(shù)采用優(yōu)先數(shù)，尺寸規(guī)格整齊，零件通用性和互換性強(qiáng)，系列容易擴(kuò)充和花樣翻新，利于組織批量生產(chǎn)和降低成本。 ③型式多樣化，變型設(shè)計(jì)多。擺脫了傳統(tǒng)的單一的底座安裝方式，增添了空心軸懸掛式、浮動(dòng)支承底座、電動(dòng)機(jī)與減速器一體式聯(lián)接，多方位安裝面等不同型式，擴(kuò)大使用范圍。 促使減速器水平提高的主要因素有： ①理論知識(shí)的日趨完善，更接近實(shí)際（如齒輪強(qiáng)度計(jì)算方法、修形技術(shù)、變形計(jì)算、優(yōu)化設(shè)計(jì)方法、齒根圓滑過渡、新結(jié)構(gòu)等）。 ②采用好的材料，普遍采用各種合金鋼鍛件，材料和熱處理質(zhì)量控制水平提高。 ③結(jié)構(gòu)設(shè)計(jì)更合理。 ④加工精度提高到ISO5－6級(jí)。 ⑤軸承質(zhì)量和壽命提高。 ⑥潤(rùn)滑油質(zhì)量提高。 自20世紀(jì)60年代以來，我國(guó)先后制訂了JB1130－70《圓柱齒輪減速器》等一批通用減速器的標(biāo)淮，除主機(jī)廠自制配套使用外，還形成了一批減速器專業(yè)生產(chǎn)廠。目前，全國(guó)生產(chǎn)減速器的企業(yè)有數(shù)百家，年產(chǎn)通用減速器25萬臺(tái)左右，對(duì)發(fā)展我國(guó)的機(jī)械產(chǎn)品作出了貢獻(xiàn)。 20世紀(jì)60年代的減速器大多是參照蘇聯(lián)20世紀(jì)40－50年代的技術(shù)制造的，后來雖有所發(fā)展，但限于當(dāng)時(shí)的設(shè)計(jì)、工藝水平及裝備條件，其總體水平與國(guó)際水平有較大差距。 改革開放以來，我國(guó)引進(jìn)一批加工裝備，通過引進(jìn)、消化、吸收國(guó)外技術(shù)和科研攻關(guān)，逐步掌握了各種高速和低速重載齒輪裝置的設(shè)計(jì)制造技術(shù)。材料和熱處理質(zhì)量及齒輪加工精度均有較大提高，通用圓柱齒輪的制造精度可從JB179－60的8－9級(jí)提高到GB10095－88的6級(jí)，高速齒輪的制造精度可穩(wěn)定在4－5級(jí)。部分減速器采用硬齒面后，體積和質(zhì)量明顯減小，承載能力、使用壽命、傳動(dòng)效率有了較大的提高，對(duì)節(jié)能和提高主機(jī)的總體水平起到很大的作用。 我國(guó)自行設(shè)計(jì)制造的高速齒輪減（增）速器的功率已達(dá)42000kW ，齒輪圓周速度達(dá)150m/s以上。但是，我國(guó)大多數(shù)減速器的技術(shù)水平還不高，老產(chǎn)品不可能立即被取代，新老產(chǎn)品并存過渡會(huì)經(jīng)歷一段較長(zhǎng)的時(shí)間。[編輯本段]減速器的設(shè)計(jì)程序 一、設(shè)計(jì)的原始資料和數(shù)據(jù) 1、原動(dòng)機(jī)的類型、規(guī)格、轉(zhuǎn)速、功率（或轉(zhuǎn)矩）、啟動(dòng)特性、短時(shí)過載能力、轉(zhuǎn)動(dòng)慣量等。 2、工作機(jī)械的類型、規(guī)格、用途、轉(zhuǎn)速、功率（或轉(zhuǎn)矩）。工作制度：恒定載荷或變載荷，變載荷的載荷圖；啟、制動(dòng)與短時(shí)過載轉(zhuǎn)矩，啟動(dòng)頻率；沖擊和振動(dòng)程度；旋轉(zhuǎn)方向等。 3、原動(dòng)機(jī) 作機(jī)與減速器的聯(lián)接方式，軸伸是否有徑向力及軸向力。 4、安裝型式（減速器與原動(dòng)機(jī)、工作機(jī)的相對(duì)位置、立式、臥式）。 5、傳動(dòng)比及其允許誤差。 6、對(duì)尺寸及重量的要求。 7、對(duì)使用壽命、安全程度和性的要求。 8、環(huán)境溫度、灰塵濃度、氣流速度和酸堿度等環(huán)境條件；潤(rùn)滑與冷卻條件（是否有循環(huán)水、潤(rùn)滑站）以及對(duì)振動(dòng)、噪聲的限制。 9、對(duì)操作、控制的要求。 10、材料、毛坯、標(biāo)準(zhǔn)件來源和庫存情況。 11、制造廠的制造能力。 12、對(duì)批量、成本和價(jià)格的要求。 13、交貨期限。 上述前四條是必備條件，其他方面可按常規(guī)設(shè)計(jì)，例如設(shè)計(jì)壽命一般為!"年。用于重要場(chǎng)合時(shí)，性應(yīng)較高等。 二、選定減速器的類型和安裝型式 三、初定各項(xiàng)工藝方法及參數(shù) 選定性能水平，初定齒輪及主要機(jī)件的材料、熱處理工藝、精加工方法、潤(rùn)滑方式及潤(rùn)滑油品。 四、確定傳動(dòng)級(jí)數(shù) 按總傳動(dòng)比，確定傳動(dòng)的級(jí)數(shù)和各級(jí)的傳動(dòng)比。 五、初定幾何參數(shù) 初算齒輪傳動(dòng)中心距（或節(jié)圓直徑）、模數(shù)及其他幾何參數(shù)。 六、整體方案設(shè)計(jì) 確定減速器的結(jié)構(gòu)、軸的尺寸、跨距及軸承型號(hào)等。 七、校校 校核齒輪、軸、鍵等負(fù)載件的強(qiáng)度，計(jì)算軸承壽命。 八、潤(rùn)滑冷卻計(jì)算 九、確定減速器的附件 十、確定齒輪滲碳深度 必要時(shí)還要進(jìn)行齒形及齒向修形量等工藝數(shù)據(jù)的計(jì)算。 十一、繪制施工圖 在設(shè)計(jì)中應(yīng)貫徹國(guó)家和行業(yè)的有關(guān)標(biāo)準(zhǔn)。[編輯本段]減速機(jī)的檢查和維護(hù) 不同的潤(rùn)滑油禁止相互混合使用。油位螺塞、放油螺塞和通氣器的位置由安裝位置決定。它們的相關(guān)位置可參考減速機(jī)的安裝位置圖來確定。 油位的檢查 ? 切斷電源，防止觸電！等待減速機(jī)冷卻！ ? 移去油位螺塞檢查油是否充滿。 ? 安裝油位螺塞。 油的檢查 ? 切斷電源，防止觸電！等待減速機(jī)冷卻！ ? 打開放油螺塞，取油樣。 ? 檢查油的粘度指數(shù) ——如果油明顯渾濁，建議盡快更換。 ? 對(duì)于帶油位螺塞的減速機(jī) ——檢查油位，是否合格 ——安裝油位螺塞 油的更換 冷卻后油的粘度增大放油困難，減速機(jī)應(yīng)在運(yùn)行溫度下?lián)Q油。 ? 切斷電源，防止觸電！等待減速機(jī)冷卻下來無燃燒危險(xiǎn)為止！ 注意：換油時(shí)減速機(jī)仍應(yīng)保持溫?zé)帷? ? 在放油螺塞下面放一個(gè)接油盤。 ? 打開油位螺塞、通氣器和放油螺塞。 ? 將油全部排除。 ? 裝上放油螺塞。 ? 注入同牌號(hào)的新油。 ? 油量應(yīng)與安裝位置一致。 ? 在油位螺塞處檢查油位。 ? 擰緊油位螺塞及通氣器。盡量選用接近理想減速比： 減速比=伺服馬達(dá)轉(zhuǎn)速/減速機(jī)出力軸轉(zhuǎn)速 扭力計(jì)算： 對(duì)減速機(jī)的壽命而言,扭力計(jì)算非常重要,并且要注意加速度的最大轉(zhuǎn)矩值（TP）,是否超過減速機(jī)之最大負(fù)載扭力. 適用功率通常為市面上的伺服機(jī)種的適用功率,減速機(jī)的適用性很高,工作系數(shù)都能維持在1.2以上,但在選用上也可以以自己的需要來決定: 要點(diǎn)有二: A.選用伺服電機(jī)的出力軸徑不能大于表格上最大使用軸徑. B.若經(jīng)扭力計(jì)算工作,轉(zhuǎn)速可以滿足平常運(yùn)轉(zhuǎn),但在伺服全額輸出時(shí),有不足現(xiàn)象時(shí),我們可以在電機(jī)側(cè)之驅(qū)動(dòng)器,做限流控制,或在機(jī)械軸上做扭力保護(hù),這是很必要的。
武漢澤林自動(dòng)化科技有限責(zé)任公司為日本電產(chǎn)新寶廠家授權(quán)代理商，代理新寶減速機(jī)ERK型（帶底座臥式）及ERK-V型（帶法蘭立式）精度型標(biāo)準(zhǔn)型。
| 產(chǎn)品類型Product type | 適用介質(zhì)Applicable medium | 適用溫度/℃Applicable temperature |
| NKZ41HNKZ44HNKZ944H-C型NKZ941H | 水、蒸汽、空氣Water，Steam，Air | ≤425 |
| NKZ41H(Y)NKZ44H(Y)-I型NKZ944H(Y)-I型NKZ941H(Y) | 水、蒸汽、空氣Water，Steam，Air | ≤550 |
主要零件材料 Materials for main parts
| 零件名稱Part name | 閥體、閥蓋Body Bonnet | 閥桿Stem | 閘板Wedge gate | 密封面Sealing face | 閥桿螺母Yokenut | 填料Parking | 緊固件Fastener |
| NKZ44HNKZ41H-C型NKZ944H-C型NKZ941H | WCB | 2Crl3 | 25 | H：合金鋼H：alloy steel | 鋁 |
flexlink系列柔性輸送板鏈無論是在水平面還是垂直面都能勝任急速的轉(zhuǎn)彎，同時(shí)摩擦力和噪音小；
flexlink系列柔性輸送板鏈廣泛應(yīng)用于醫(yī)藥、機(jī)械零件、食品和飲料工業(yè)、化妝品行業(yè)、電機(jī)制造業(yè)、汽車零件、軸承制造業(yè)、電子等
1、flexlink輸送鏈
| 鏈板 寬度 | 基本 型號(hào) | 平板鏈 | 通用鏈 | 鋼頂鏈 | 楔形鏈 | 滾珠鏈 | 彈性楔鏈 |
| 44 | CS | CSTP5 |
|
| CSTF5 |
|
|
| 63 | CL | CLTP5 |
| CLTP5TF | CLTF5 | CLTR5 | CLTE5 |
| 83 | CM | CMTP5 | CMTF5U | CMTP5 | CMTF5 | CMTR5 | CMTE5C |
| 103 | CH | CHTP5 |
| CHTP5TF | CHTF5 | CHTR5 |
|
| 175 | CB | CBTP3A |
|
| CBTL |
|
|
2、flexlink輸送粱與連接件
| 鏈板寬度 | 輸送粱寬度 | 輸送粱厚度 | 輸送粱型號(hào) | 連接件型號(hào) |
| 43 | 45 | 64 | CSCB3 |
|
| 63 | 65 | 64.2 | CLCB3 | CLCC160 |
| 83 | 85 | 75 | CMCB3 | CMCC160 |
| 103 | 105 | 75 | CHCB3 | CHCC160 |
3、flexlink柔性輸送線性能參數(shù)
（1）單個(gè)驅(qū)動(dòng)頭的輸送長(zhǎng)度：25M
（2）工作溫度：-20℃～+60℃
（3）輸送速度：50M/min
（4）鏈節(jié)距：38.1mm
（5）最小轉(zhuǎn)彎半徑：R150mm （6）抗拉強(qiáng)度：4000N （7）鏈板重量：0.7KG/M 4、flexlink柔性輸送線特點(diǎn)
（1）潔凈：整線由白色高強(qiáng)度工程塑料鏈板與表面經(jīng)陽極氧化的鋁合金異型材組裝而成。鏈板為純白色，設(shè)備運(yùn)行無需潤(rùn)滑，無須定期保養(yǎng)，不會(huì)被腐蝕，符合GMP規(guī)范。
（2）安靜：設(shè)備運(yùn)行聲音＜30Db。
（3）靈巧：由可任意拆裝的組件裝配成具有彈性的傳送系統(tǒng)，配合不同半徑的垂直和水平彎道，可以在任意的3D空間內(nèi)輸送產(chǎn)品，并可隨時(shí)隨地按照你的生產(chǎn)現(xiàn)狀進(jìn)行調(diào)整？梢酝瓿伤健⑴榔、垂直、轉(zhuǎn)彎、螺旋、夾持、懸掛、翻轉(zhuǎn)、旋轉(zhuǎn)等輸送形式。
（4）簡(jiǎn)便：整線安裝無需任何特殊工具，只需常用手工工具，即可由單人完成基本拆裝工作。
（5）穩(wěn)定：穩(wěn)定的性能可保證被送的產(chǎn)品在輸送的過程中不發(fā)生翻倒和滑落。•Solid state design is safer than solenoid testers.•Low impedance to reduce false readings.•Indicates voltage without battery.•Voltage indicated by lights, vibration ａnd tone.•With continuity beeper ａnd bright work light. •固態(tài)設(shè)計(jì)，比電磁測(cè)試儀安全。•低阻抗，以減少錯(cuò)誤的讀數(shù)。•指示，不帶電池的電壓。•電壓表示燈，振動(dòng)和音。•連續(xù)性蜂鳴器和明亮的工作燈。
| 技術(shù)參數(shù)：德國(guó)凱馳Karcher（廣州博勵(lì)）全自動(dòng)洗地機(jī)吸干機(jī) | |
| 洗地寬度 毫米 | 800 |
| 吸水寬度 毫米 | 990 |
| 凈/污水箱 升 | 120/120 |
| 洗地效率 平米/小時(shí) | 3200 |
| 刷子轉(zhuǎn)速 轉(zhuǎn)/分 | 200 |
| 刷子觸地壓力（可調(diào)） | 36 克/平方厘米 |
| 電機(jī)功率 瓦 | 1660 |
| 重量 公斤 | 170 |
電線桿裂縫寬度測(cè)試儀主要技術(shù)參數(shù)：
主機(jī)參數(shù)屏幕尺寸：5英寸 液晶分辨率：160×128 體積：195×140×45mm 重量：0.8kg主機(jī)外殼防水、防塵、防震測(cè)量范圍0.01 ~ 2.00mm讀數(shù)精度0.005mm放大倍數(shù)40倍最小分度0.02mm供電方式內(nèi)置鋰電池連接線長(zhǎng)2.5m主機(jī)重量0.8kg工作環(huán)境溫度-10℃～40℃包裝規(guī)格材質(zhì)：工程塑料體積：×315×170mm 重量：5kg
Rat Interleukin 10（IL-10）
ELISA Kit
Catalog No. CSB-E04595r
（96T）
l This immunoassay kit allows for the in vitro quantitative determination of rat IL-10 concentrations in serum, plasma ａnd Tissue Homogenates.
l Expiration date six months from the date of manufacture
l FOR RESEARCH USE ONLY. NOT FOR USE IN DIAGNOSTIC PROCEDURES.
PRINCIPLE OF THE ASSAY
The microtiter plate provided in this kit has been pre-coated with an antibody specific to IL-10. Standards or samples are then added to the appropriate microtiter plate wells with a biotin-conjugated polyclonal antibody preparation specific for IL-10 and Avidin conjugated to Horseradish Peroxidase （HRP） is added to each microplate well ａnd incubated. Then a TMB （3,3＇,5,5＇ tetramethyl-benzidine） substrate solution is added to each well. Only those wells that contain IL-10, biotin-conjugated antibody ａnd enzyme-conjugated Avidin will exhibit a change in color. The enzyme-substrate reaction is terminated by the addition of a sulphuric acid solution ａnd the color change is measured spectrophotometrically at a wavelength of 450 nm ± 2 nm. The concentration of IL-10 in the samples is then determined by comparing the O.D. of the samples to the standard curve.
DETECTION RANGE
3.12 pg/ml-200 pg/ml. The standard curve concentrations used for the ELISA’s were 200 pg/ml, 100 pg/ml, 50 pg/ml, 25 pg/ml, 12.5 pg/ml, 6.25 pg/ml, 3.12 pg/ml.
SPECIFICITY
This assay recognizes rat IL-10. No significant cross-reactivity or interference was observed.
SENSITIVITY
The minimum detectable dose of rat IL-10 is typically less than 0.78 pg/ml.
The sensitivity of this assay, or Lower Limit of Detection （LLD） was defined as the lowest protein concentration that could be differentiated from zero.
MATERIALS PROVIDED
| Reagent | Quantity |
| Assay plate | 1 |
| Standard | 2 |
| Sample Diluent | 1 x 20 ml |
| Biotin-antibody Diluent | 1 x 10 ml |
| HRP-avidin Diluent | 1 x 10 ml |
| Biotin-antibody | 1 x 120μl |
| HRP-avidin | 1 x 120μl |
| Wash Buffer | 1 x 20 ml （25×concentrate） |
| TMB Substrate | 1 x 10 ml |
| Stop Solution | 1 x 10 ml |
STORAGE
1. Unopened test kits should be stored at 2-8°C upon receipt ａnd the microtiter plate should be kept in a sealed bag. The test kit may be used throughout the expiration date of the kit, provided it is stored as prescribed above. Refer to the package label for the expiration date.
2. Opened test plate should be stored at 2-8°C in the aluminum foil bag with desiccants to minimize exposure to damp air. The kits will remain stable until the expiring date shown, provided it is stored as prescribed above.
3. A microtiter plate reader with a bandwidth of 10 nm or less ａnd an optical density range of 0-3 OD or greater at 450nm wavelength is acceptable for use in absorbance measurement.
REAGENT PREPARATION
Bring all reagents to room temperature before use.
1. Wash Buffer If crystals have formed in the concentrate, warm up to room temperature ａnd mix gently until the crystals have completely dissolved. Dilute 20 ml of Wash Buffer Concentrate into deionized or distilled water to prepare 500 ml of Wash Buffer.
2. Standard Centrifuge the standard vial at 6000-10000rpm for 30s. Reconstitute the Standard with 1.0 ml of Sample Diluent. This reconstitution produces a stock solution of 200 pg/ml. Allow the standard to sit for a minimum of 15 minutes with gentle agitation prior to making serial dilutions. The undiluted standard serves as the high standard （200 pg/ml）. The Sample Diluent serves as the zero standard （0 pg/ml）. Prepare fresh for each assay. Use within 4 hours ａnd discard after use.
3. Biotin-antibody Centrifuge the vial before opening. Dilute to the working concentration using Biotin-antibody Diluent（1:100）, respectively.
4. HRP-avidin Centrifuge the vial before opening. Dilute to the working concentration using HRP-avidin Diluent（1:100）, respectively.
Precaution: The Stop Solution provided with this kit is an acid solution. Wear eye, hand, face, ａnd clothing protection when using this material.
OTHER SUPPLIES REQUIRED
— Microplate reader capable of measuring absorbance at 450 nm, with the correction wavelength set at 540 nm or 570 nm.
— Pipettes ａnd pipette tips.
— Deionized or distilled water.
— Squirt bottle, manifold dispenser, or automated microplate washer.
— An incubator which can provide stable incubation conditions up to 37°C±0.5°C.
SAMPLE COLLECTION AND STORAGE
l Serum Use a serum separator tube （SST） ａnd allow samples to clot for 30 minutes before centrifugation for 15 minutes at 1000 g. Remove serum ａnd assay immediately or aliquot and store samples at -20°C. Centrifuge the sample again after thawing before the assay. Avoid repeated freeze-thaw cycles.
l Plasma Collect plasma using citrate, EDTA, or heparin as an anticoagulant. Centrifuge for 15 minutes at 1000 g within 30 minutes of collection. Assay immediately or aliquot ａnd store samples at -20°C. Centrifuge the sample again after thawing before the assay. Avoid repeated freeze-thaw cycles.
l Tissue Homogenates 100mg tissue was rinsed with 1X PBS, homogenized in 1 mL of 1X PBS ａnd stored overnight at -20° C. After two freeze-thaw cycles were performed to break the cell membranes, the homogenates were centrifuged for 5 minutes at 5000 x g, 2 - 8°C. The supernate was assayed ａnd removed immediately. Alternatively, aliquot ａnd store samples at -20°C or -80℃. Centrifuge the sample again after thawing before the assay. Avoid repeated freeze-thaw cycles.
Note: Grossly hemolyzed samples are not suitable for use in this assay.
ASSAY PROCEDURE
Bring all reagents ａnd samples to room temperature before use. It is recommended that all samples, standards, ａnd controls be assayed in duplicate. All the reagents should be added directly to the liquid level in the well. The pipette should avoid contacting the inner wall of the well.
1. Add 100μl of Standard, Blank, or Sample per well. Cover with the adhesive strip. Incubate for 2 hours at 37°C.
2. Remove the liquid of each well, don’t wash.
3. Add 100μl of Biotin-antibody working solution to each well. Incubate for 1 hour at 37°C. Biotin-antibody working solution may appear cloudy. Warm up to room temperature ａnd mix gently until solution appears uniform.
4. Aspirate each well ａnd wash, repeating the process three times for a total of three washes. Wash: Fill each well with Wash Buffer （200μl） ａnd let it stand for 2 minutes, then remove the liquid by flicking the plate over a sink. The remaining drops are removed by patting the plate on a paper towel. Complete removal of liquid at each step is essential to good performance.
5. Add 100μl of HRP-avidin working solution to each well. Cover the microtiter plate with a new adhesive strip. Incubate for 1 hour at 37°C.
6. Repeat the aspiration ａnd wash five times as step 4.
7. Add 90μl of TMB Substrate to each well. Incubate for 10-30 minutes at 37°C. Keeping the plate away from drafts ａnd other temperature fluctuations in the dark.
8. Add 50μl of Stop Solution to each well when the first four wells containing the highest concentration of standards develop obvious blue color. If color change does not appear uniform, gently tap the plate to ensure thorough mixing.
9. Determine the optical density of each well within 30 minutes, using a microplate reader set to 450 nm.
CALCULATION OF RESULTS
Using the professional soft "Curve Exert 1.3" to make a standard curve is recommended, which can be downloaded from our web.
Average the duplicate readings for each standard, control, ａnd sample ａnd subtract the average zero standard optical density. Create a standard curve by reducing the data using computer software capable of generating a four parameter logistic （4-PL） curve-fit. As an alternative, construct a standard curve by plotting the mean absorbance for each standard on the x-axis against the concentration on the y-axis ａnd draw a best fit curve through the points on the graph. The data may be linearized by plotting the log of the IL-10 concentrations versus the log of the O.D. ａnd the best fit line can be determined by regression analysis. This procedure will produce an adequate but less precise fit of the data. If samples have been diluted, the concentration read from the standard curve must be multiplied by the dilution factor.
LIMITATIONS OF THE PROCEDURE
— The kit should not be used beyond the expiration date on the kit label.
— Do not mix or substitute reagents with those from other lots or sources.
— It is important that the Standard Diluent selected for the standard curve be consistent with the samples being assayed.
— If samples generate values higher than the highest standard, dilute the samples with the appropriate Standard Diluent ａnd repeat the assay.
— Any variation in Standard Diluent, operator, pipetting technique, washing technique, incubation time or temperature, ａnd kit age can cause variation in binding.
— This assay is designed to eliminate interference by soluble receptors, binding proteins, and other factors present in biological samples. Until all factors have been tested in the Immunoassay, the possibility of interference cannot be excluded.
TECHNICAL HINTS
— Centrifuge vials before opening to collect contents.
— When mixing or reconstituting protein solutions, always avoid foaming.
— To avoid cross-contamination, change pipette tips between additions of each standard level, between sample additions, ａnd between reagent additions. Also, use separate reservoirs for each reagent.
— When using an automated plate washer, adding a 30 second soak period following the addition of wash buffer, and/or rotating the plate 180 degrees between wash steps may improve assay precision.
— To ensure accurate results, proper adhesion of plate sealers during incubation steps is necessary.
— Substrate Solution should remain colorless or light blue until added to the plate. Keep Substrate Solution protected from light. Substrate Solution should change from colorless or light blue to gradations of blue.
— Stop Solution should be added to the plate in the same order as the Substrate Solution. The color developed in the wells will turn from blue to yellow upon addition of the Stop Solution. Wells that are green in color indicate that the Stop Solution has not mixed thoroughly with the Substrate Solution.