![SMOBIO/[QP5210] Q-PAGE™ TGN Precast Gel (Midi, 12 wells, 10%), 10 gels/Midi, 12 wells, 10%), 10 gels</span>
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Description
Q-PAGE™ TGN (Tris-Glycine Novel) Precast Gels are ready-to-use acrylamide gels for SDS-PAGE running in Tris-Glycine buffer system. With unique formula, Q-PAGE™ TGN Precast Gels perform enhanced speed, better separation, and longer shelf life as compared with conventional Laemmli Tris-HCl gels. The protein migration patterns in Q-PAGE™ TGN series, however, are similar with typical Laemmli Tris-HCl gels, and thus Q-PAGE™ TGN Precast Gels are compatible to traditional SDS-PAGE and subsequent analyses.
Q-PAGE™ TGN Precast Gels are available in gradient (4 to 15%) and fixed (10%) concentrations of polyacrylamide in 12- and 15-well formats. Two available cassette sizes, Mini (10 x 8.3 cm) and Midi (10 x 10 cm), are compatible with most popular protein electrophoresis systems. Q-PAGE™ Mini (QP4XXX) Gels are suitable for Bio-Rad® and other systems. Q-PAGE™ Midi (QP5XXX) Gels are suitable for Invitrogen® XCell SureLock® Mini-Cell, Invitrogen® Mini Gel Tank, Hoefer SE260, and other systems.
Key Features
User-friendly gel cassette:
Numbered and framed wells for sample loading
Labeled warning sign and green tape as reminder
Enhanced gel performance:
Enhanced gel electrophoresis speed
Better band separation
Stable for shipping at ambient temperature
Easy compatibility:
Available as homogeneous and adjusted gradient gels for a wide range of protein separation.
Compatible with most popular protein electrophoresis systems
Storage and stability
Store Q-PAGE™ Precast Gels at 4°C for periods up to 12 months.
Do not freeze Q-PAGE™ Precast Gels. Remove tape and comb before electrophoresis.
Quick running, clear bands
Q-PAGE™ TGN Precast Gel can separate protein in 19 minutes using 300 V.
QP5210 Specifications
Gel | TGN(Tris-Glycine-Novel) | |
Buffersystems | Tris-Glycine (Laemmli) | |
Features | Quickrunning, clear bands | |
Cassettesize | Midi Gel (10 X 10 cm) | |
Geldimensions | 8.1 x 8.1 x0.1 cm (W x L xthickness) cm | |
Electrophoresissystem | Mini Gel Tank XCell SureLock, Hoefer SE260 | |
Well format& Capacity | 12 wells, 40 μl/well | |
Gelpercentage | 10 % | |
Accessorytray | Productiondescription Tip card Gel remover Cassetteopener | |
Manual
Manual_Q-PAGE™ TGN Precast Gel, Midi
SDS
SDS_Q-PAGE™ Precast Gel
Migration pattern
Setting Up and Running Q-PAGE™ Midi Precast Gel
Removing Q-PAGE Midi Gel from cassette
Setting up gel/membrane sandwich for Western transfer
Recommendations/Tips for Gel Running
1. Remove comb and tape before adaption. 2. Use fresh 1X running buffer for the inner cathode chamber. 3. Rinse the wells before sample loading. 4. Try 200 V first, and optimize the voltage and running time if needed. Do not set voltage lower than 100 V.
Sample Preparation for SDS-PAGE
1. Mix protein sample with 2X sample buffer.
2. Heat the diluted samples at 95°C for 5 min or at 70°C for 10 min.
3. Cool the diluted samples to 4°C and spin down the water condensed on tube surface. (If there is high viscosity part at bottom of tube, transfer supernatant to a new tube.)
Prepare Q-PAGE™ for Sample Loading
1.Open the blister tray of Q-PAGE™ Precast Gel.
2.Briefly rinse the gel cassette with ddH2O.
3.Remove tape and comb; avoid squeezing the gel.
4.Adapt Q-PAGE™ to electrophoresis system; instruction are provided below. (Invitrogen® Mini Gel Tank is recommended.)
5.Use a pipette to gently wash the wells with running buffer to remove residual storage buffer.
6.Fill the wells with running buffer prior to sample loading.
7.Load samples and pre-stained protein marker into numbered wells.
8.Fill both inner and outer chambers with running buffer to the highest level. Ensure gel wells are completely covered.
Power Setting for Running Q-PAGE™
Optimize the voltage and running time if needed.
| 150 V | 200 V*2 | 250 V*3 | 300 V*3 |
Running Time*1 | 50-70 mins | 35-55 mins | 25-40 mins | 15-30 mins |
Expected Current Initial (per gel) Final (per gel) |
35-45 mA 10-20 mA |
45-55 mA 20-25 mA |
75-85 mA 40-45 mA |
100-110 mA 60-70 mA |
Expected temperature | 25-30°C | 25-30 °C | 25-35°C | 30-40°C |
*1 Set voltage higher than 100 V is recommended.
*2 Try 200 V first, and optimize the voltage and running time if needed
*3 For higher voltage conditions, please use fresh running buffer for inner and outer chambers
*4 Running time varies depending on gel percentage, running buffer, temperature, and power supply.
Remove Q-PAGE™ Gel from Cassette
Open cassette immediately after electrophoresis. Avoid gel drying.
1.Insert the cassette opener into corners of cassette.
2.Sequentially pry the opener to separate the two plates.
3.Gently pull up notched plate and let gel stay on the front plate.
4.Use cassette opener to push through the slot in the cassette.
5.Carefully detach the gel from the bottom of gel.
- Avoid diagonally peeling the gel from the corner.
- If necessary, cut well separators with gel remover.
6.Gently remove the gel for further staining or Western blotting.
Gel Staining
Proteins separated using Q-PAGE™ Precast Gels can be further stained with most popular staining reagents, such as Coomassie dyes (R-250 or G-250), Silver-stain solution,
and FluoroStain™ Protein Fluorescent Staining Dye. (Cat. No. PS1000)
Transferring Protein from Q-PAGE™ to Blotting Membrane
1. After protein separation using Q-PAGE™, gently detach QPAGE™ from cassette and then equilibrate the gel in transfer buffer.
2. Pre-soak blotting membrane and filter papers in transfer buffer.
*Activate PVDF membrane in methanol before soaking in transfer buffer.
**Prepare 6 filter papers for one gel/membrane sandwich.
3. Assemble transfer sandwich by orientating cathode, sponge, filter papers, gel, membrane, filter papers, sponge, and anode. The protein goes to the direction of cathode to anode.
4. Carefully move roller over the gel/membrane to remove air bubbles and excess buffer until complete contact is established.
5. Insert transfer cassette into transfer module. Notice that black side of cassette should be next to black side of module.
6. Fill transfer tank with pre-cooled transfer buffer to the highest water level.
7. Set constant voltage at 100 V. Transfer for 90 minutes at low temperature condition. Pre-stained protein marker should be visible on the membrane after transfer is completed.
Transfer of proteins to the membrane can be checked using Ponceau S staining before blocking step.
Supplemental Information for Using Q-PAGE™ Precast Gel
Adapting Q-PAGE™ Midi Precast Gels to Invitrogen Mini Gel Tank Electrophoresis System
1. Place the Q-PAGE Midi Precast Gels with notched plate facing toward yourself. No extra adapter is needed.
2. Seat the gels on the bottom of Mini Gel Tank and close the cassette clamp.
3. Fill chambers with running buffer to the level of the fill line. Ensure gel wells are completely covered.
Adapting Q-PAGE™ Midi Precast Gels to other electrophoresis system, please follow the manufacturer’s instruction.
Buffer recipes
2X sample buffer with reducing agent
62.5 mM Tris-HCl pH 6.8, 2% SDS, 25% (v/v) glycerol, 0.01% bromophenol blue, 5% β-mercaptoethanol or 100 mM DTT (added fresh)
10X Tris-Glycine running buffer
30.0 g Tris base, 144.0 g Glycine, 10.0 g SDS. Bring up the volume to 1 L with ddH2O.
1X running buffer
Dilute 100 ml 10X running buffer with 900 ml ddH2O.
10X transfer buffer
30.0 g Tris base, 144.0 g Glycine. Bring up the volume to 1 L with ddH2O.
1X transfer buffer
*Cool 1X transfer buffer to 4°C before using.
Dilute 100 ml 10X transfer buffer with 200 ml methanol and 700 ml ddH2O.
**Add SDS to 0.1% to promote transfer of high molecular weight proteins.
Troubleshooting Guidelines | ||
Problem | Possible Cause | Suggested Solution |
Well deformation | Pull one side of comb out of cassette. | Smoothly pull the comb straight out of the cassette. |
Bubbles between gel and cassette | Gel has been frozen or stored at wrong temperature. | Store Q-PAGE Precast Gels at 4°C. |
Buffer leaking from the inner chamber | Untight assembly of gels to the electrode modules | Reassemble Q-PAGE gels into the electrodemodules. Fill outer chamber with 1X running buffer to thehighest level. |
Samples do not sink into the wells. | Residual gel storage buffer in the wells | Rinse the gel wells with ddH2O or 1X running bufferbefore loading. |
Insufficient sample buffer | Use more sample buffer to prepare samples. | |
Current is zero and sample do not migrate into gel | Tape at bottom of gel not removed | Remove tape |
Gels run faster or more slowly than expected. | Incorrect running buffer | Check buffer composition. Use fresh 1X running buffer for inner chamber. |
Crooked bands at middle or bottom of gel | Gel has been frozen or stored at wrong temperature. | Store Q-PAGE Precast Gels at 4°C. |
Incorrect running buffer | Check buffer composition. Use fresh 1X running buffer for inner chamber. | |
Band pattern curves toward one or both sides of gel. | Buffer leaking from the inner chamber | Check assembly of gels into the electrode modules. |
Excessive heating of gel | Check buffer composition. Or dilute running bufferto 0.5-0.75X. Do not exceed recommended running conditions. | |
Insufficient buffer in inner or outer buffer chamber | Fill inner and outer chambers to completely covergel wells. | |
Poor resolution or fuzzy bands | Excessive heating of gel | Check buffer composition. Do not exceed recommended running conditions. |
Incorrect running buffer | Check buffer composition. | |
Bands are missing on the membrane after Westerntransferring. | Proteins move in the wrong direction | Check the order of gel/membrane sandwich assembly,the direction of transfer cassette in transfer modules, and the polarity ofconnections to power supply. |
Swirls or missing bands; bands trail off in multipledirections on the membrane after Western transferring. | Contact between the membrane and the gel was poor;Air bubbles or excess buffer remains between the blotting membrane andthe gel. | Use thicker/more filter paper in the gel/membranesandwich Remove air bubbles and excess buffer betweengel and membrane by carefully moving the roller over the membrane. |
Apparent molecular sizes of prestained proteinmarkers are different as indicated. | Prestained protein markers used have not beencalibrated for use with Q-PAGE gels. Dyes for staining protein markers affect themigration patterns of prestained proteins in different buffer systems. | Calibrate prestained protein markers againstunstained proteins of known size or use SMOBIO’s ExcelBand™ Protein Markers. |
Q-PAGE™ Precast Gel
Gel Type | Bis-Tris | TGN (Tris-Glycine-Novel) | ||||||
Buffer systems | MOPS and MES | Tris-Glycine (Laemmli) | ||||||
Features | Clear and sharp bands, high resolution | Quick running, clear bands | ||||||
Cassette size | Mini Gel(10 x 8.3 cm) | Midi Gel(10 X 10 cm) | Mini Gel(10 x 8.3 cm) | Midi Gel(10 X 10 cm) | ||||
Electrophoresis system | Bio-Rad systems | Mini Gel Tank Xcell SureLock, Hoefer SE260 | Bio-Rad systems | Mini Gel Tank Xcell SureLock, Hoefer SE260 | ||||
Well format & Capacity | 12 wells, 25 μl/well | 15 wells, 22 μl/well | 12 wells, 40 μl/well | 15 wells, 28 μl/well | 12 wells, 25 μl/well | 15 wells, 22 μl/well | 12 wells, 40 μl/well | 15 wells,28 μl/well |
Gel percentage/ Cat. No. | 8% | 8% | 8% | 8% | 10% | 10% | 10% | 10% |
QP2110 | QP2120 | QP3110 | QP3120 | QP4210 | QP4220 | QP5210 | QP5220 | |
12% | 12% | 12% | 12% | 4-15% | 4-15% | 4-15% | 4-15% | |
QP2310 | QP2320 | QP3310 | QP3320 | QP4510 | QP4520 | QP5510 | QP5520 | |
4-12% | 4-12% | 4-12% | 4-12% |
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QP2510 | QP2520 | QP3510 | QP3520 |
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ExcelBand™ Protein Markers
Ready-to-use— premixed with a loading buffer for direct loading, no need to boil
Broad range— 310 kDa to 5 kDa
Pre-stained bands — for monitoring protein separation during electrophoresis and Western blotting transferring efficiency on membrane
Enhanced bands— for quick reference
YesBlot™ Western Marker I
Ready-to-use — no need of mixing or heating before sample loading
Direct visualization — 10 IgG-binding proteins for direct visualization on Western blots
Pre-stained bands — 4 pre-stained proteins for monitoring protein separation during electrophoresis and Western blotting transferring efficiency on membrane
Wide range — 10 clear bands from 15 to 200 kDa for size estimation
Quick reference — two enhanced bands (30 and 80 kDa)
FluoroStain™ Protein Fluorescent Staining Dye
Compatible to MASS analysis — compatible to the analysis of mass spectra, such as LC-MS/MS, MALDI-TOF, and etc.
High sensitivity — detection level achieve ~3 ng, similar to silver staining
Substitution of the Coomassie Blue protein staining method
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质粒转染一般是说DNA转染，用常用的sinofection等转染试剂就可以；
小干扰RNA（Small interfering RNA；siRNA）有时称为短干扰RNA（short interfering RNA）或沉默RNA（silencing RNA），是一个长20到25个核苷酸的双股RNA，这种RNA的转染需要用专门的RNA转染试剂。
如题，PolyplusTransfection转染试剂在中国区的代理商有哪些？求推荐1-2个靠谱的，谢谢！
想用RNAiMAX或者lipo2000转染siRNA，但是看了下转染试剂的说明书和锐博的siRNA说明书，觉得分别对siRNA的用量描述差别挺大的呀，不知道到底该参考哪个呢。
以24孔板为例在siRNA说明书中写到，siRNA终浓度是50nM的话，加入浓度为20μM的siRNA1.25ul，每孔体积是500ul，那这样的话，每孔最终siRNA的量是25pmol。
但是在RNAiMAX或者是lipo2000说明书中，一个写的每孔siRNA用量是5pmol，一个是500ng，这与siRNA厂家所提供的量相差也太多了吧。
到底该看哪一个呢。
ps.一旦siRNA的量和体积确定下来之后，转染试剂的量和siRNA1:1的加就可以了吗？
请各位大神解答。
1.siRNA说明书中的用量，红线圈出
2.RNAIMAX说明书中siRNA的用量。
3.lipo2000说明书中siRNA用量
对重复性要求不高的试验，一般没有问题的，只不过可能需要增加一些转染试剂用量了。
为了避免这个问题，推荐收到货之后，进行适当分装，用多少，拿出来多少最好。
但是在选择的时候，也需要注意其他的一些问题。
1、采用何种原料和抗体，是否高效、灵敏、特异
2、规范包被操作，吸附是否均匀
3、重复性、可靠性
6、是否提供技术服务
7、适用于血浆、血清、组织匀浆液、细胞培养上清液、尿液等多种类型的样本
8、可检测动物类型是否丰富
9、可检测指标是否齐全
elisa试剂盒就查下博欧特生物
使用方法：
1、 血清：操作过程中避免任何细胞刺激。使用不含热原和内毒素的试管。收集血液后，1000×g离心10分钟将血红细胞迅速小心地分离。
2、 血浆：EDTA、柠檬酸盐、肝素血浆可用于检测。1000×g离心30分钟去除颗粒。
3、 细胞上清液：1000×g离心10分钟去除颗粒和聚合物。
4、 组织匀浆：将组织加入适量生理盐水捣碎。1000×g离心10分钟，取上清液。
5、 保存：如果样品不立即使用，应将其分成小部分-70℃保存，避免反复冷冻。尽可能的不要使用溶血或高血脂血。如果血清中大量颗粒，检测前先离心或过滤。不要在37℃或更高的温度加热解冻。应在室温下解冻并确保样品均匀地充分解冻。
内毒素是革兰氏阴性菌细胞壁(cellwall)上的特有成分，主要是脂多糖中的类脂A，在细菌被裂解时被释放出来，由于其化学结构和特性，在质粒的纯化过程中很容易混入质粒DNA一同提取出来。内毒素的存在会严重的影响质粒转染细胞的效率，此外会激活造血细胞（如B细胞、巨噬细胞等）的非特异免疫反应，造成实验的假阳性，所以转染级质粒的提取纯化必须去除内毒素。
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