Salim, SELEX method is more acurate than others, and is a emerging for small molecules.

GO SELEX (Graphene Oxide SELEX) is most appropriate SELEX for screening high affinity aptamer for small molecules. It is immobilisation free SELEX, both target and NA library is free to interact, which makes it flexible assay.

There isn't a Best Method, however Recent advances have contributed to easier, quicker, and more reliable methods for the selection of aptamers for small molecules as namely conventional/Flumag and Capture/structure-switching. An often-used method is Structure-Switching or Capture SELEX, now it's dependent on the molecule which you want to select. if you need more informations contact me.

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Aptamers have a wide range of targets, and the size and solubility of the targets are different. Therefore, the application of SELEX technology to screen nucleic acid aptamers can adopt different specific operation methods. Centrifugal precipitation is often used for aptamer screening of cells, bacteria and viruses, and solid-phase adsorption and elution technology can be used for aptamer screening of soluble small molecules. Specific methods for aptamer screening include the following aspects.

Screening methods based on different fixation media. Nitrocellulose membrane filtration is a commonly used method for protein aptamer screening.

Joshi et al. designed a lateral flow chromatographic device based on nitrocellulose membrane. Combining the device with SELEX screening technology, the authors obtained a high concentration of the outer membrane protein (Omp) of Salmonella typhimurium after 7 rounds of positive screening and 3 rounds of reverse screening. Affinity aptamer 33 and 45 sequences with a detection limit of 10-40 cfu/mL. Gel column is also a common immobilization medium for aptamer screening.

Tang et al. used SELEX technology to screen aptamers with high specificity and high affinity for red bean toxin based on agarose gel resin, and their dissociation constants were as low as several nmol. In recent years, the use of microwell plates as immobilization media to screen aptamers has been widely used. Wang Lifeng et al. screened and obtained a high-affinity aptamer sequence that can specifically bind to carcinoembryonic antigen (CEA) based on microwell plate technology. Studies have shown that this aptamer plays an important role in the early diagnosis, monitoring and treatment of tumors.

Fluorescent magnetic beads SELEX (FluMag-SELEX) technology. FluMag-SELEX is a method for screening fluorescent aptamers by combining the application of magnetic beads and SELEX technology. This method requires a small amount of targets and can directly measure the amount of binding ligands through fluorescence. Kim et al. applied FluMag-SELEX technology to screen five specific aptamers of ibuprofen, a broad-spectrum anti-inflammatory drug. Three of them are specific aptamers for racemic ibuprofen, and the other two can specifically interact with racemic and meso ibuprofen. The ibuprofen aptamer has strong specificity and has no binding effect with ibuprofen analogues and oxytetracycline. Xu et al. used FluMag-SELEX technology to screen specific aptamers for PCBs, with dissociation constants at the micromolar level and good linearity in the range of 0.1 to 100 ng/mL.

Capillary Electrophoresis SELEX Technology (CE-SELEX). There is a certain difference in the charge-to-mass ratio between different components, resulting in a difference in the electrophoretic mobility of the substance, thereby achieving the separation of different components. CE-SELEX can realize the screening of high-affinity aptamers within 2-4 rounds, and is often used to screen macromolecular substances such as proteins, lipopolysaccharides, and polypeptides. For the first time, Yang et al. used CE-SELEX to screen the aptamers of the small molecule substance methylmorpholine. After three rounds of screening, eight high-affinity aptamers were obtained, with dissociation constants ranging from several hundred nM to several uM, two of the sequences can catalyze the metal insertion reaction of mesoporphyrin, and the catalytic strength is 1.7 times and 2 times, respectively. Aiming at the problem of dynamic dissociation equilibrium between aptamers with weaker affinity and target systems, researchers have developed non-equilibrium capillary electrophoresis (NECEEM) of equilibrium mixtures, equilibrium capillary electrophoresis of equilibrium mixtures (ECEEM) and Non-SELEX capillary electrophoresis Electrophoresis technology.

Ashley et al. used Non-SELEX technology to screen the aptamers of catalase. The authors characterized their affinity and specificity by using a fluorescence spectrometer and capillary affinity electrophoresis. The affinity of the protein is as high as 100 times, which also shows its high specificity. The aptamer can be applied to biosensor, immunoblotting and biomarker identification. Ashley et al. screened the aptamers of human leptin protein in a free three-dimensional space environment based on NECEEM and SELEX technology, and its dissociation constant was several hundred nM.

Cell SELEX (Cell-SELEX) technology. The main target substances of this method are cells, bacteria or viruses, etc. During the operation, centrifugation and precipitation methods are used to separate and remove unbound suitable ligands, and then specific aptamer sequences are obtained through thermal dissociation or enzyme digestion.

Liang et al. used Cell-SELEX to obtain 5 DNA aptamers after 35 rounds of repeated screening for living cells infected with rabies virus. Virus titer determination and real-time quantitative reverse transcription PCR experiments showed that the five aptamers screened could inhibit the replication of rabies virus, which provided a possibility for the treatment of rabies infection.

Ninomiya et al. used Cell-SELEX to obtain 12 high-affinity aptamers that specifically bind to human liver cancer cell HepG2 after 11 rounds of repeated screening, with dissociation constants ranging from 19-450 nM. The author analyzed and obtained the secondary structure shared by 12 aptamers, which is closely related to the recognition of HepG2. This method can be screened when the nature of the target is unclear and the binding site is not determined, and it does not require a complicated process for preparing the target.tamer screening of cells, bacteria and viruses, and solid-phase adsorption and elution technology can be used for aptamer screening of soluble small molecules.