Several methods are employed for the transfection of mammalian neural cells. Each method presents its advantages and complexities. There is no single method discovered yet that can be used for the transfection of each cell type. More research is being done for the discovery of even better methods that can be used for transfection in the future. Here are some common transfection methods and their complexities: 

Electrical Transfection method:

The electrical transfection method, also known as electroporation, changes the overall characteristics of the cellular membrane by the exposure of a cell to electrical voltage. This permits the entry of charged extracellular material into the cell. The external plasmid mainly enters the cytoplasm, so there is no obvious expression of this foreign plasmid in postmitotic cells such as neural cells. The transfection ability of this method is limited because of the premature voltage applied to the cells.

Nucleofection:

Nucleofection is the modified version of electroporation in which electrical voltage is applied for the direct transfer of plasmid into the nucleus. More advances in this technique have resulted in the transfection of neural cells without affecting their proliferation abilities. This is interesting in the conditions of certain diseases because the neural cells with the controlled proliferation capacities promise the repair and regeneration of the nervous system.

Single-cell electroporation:

It is also one of the advanced versions of electroporation in which a single cell (neuron) is targeted and transfected by expression plasmids. The target neuron is identified with the help of two-photon microscopy. After its identification, it is targeted by a patch pipette in vivo that contains the plasmid DNA. Stable transgene expression in the transfected neuron can be observed for days and even months. After a certain time period, the same target cell can be transfected with different plasmid constructs. This allows the multiple transgene expression in the same cell which assesses the effects of various gene expressions during neural cell differentiation. 

Ca²⁺-phosphate/DNA coprecipitation:

This is one of the best transfection techniques used for transfecting primary neural cells and cell lines. In this procedure, the DNA crystals are formed using Ca²⁺ ions in the phosphate buffer. The crystals are precipitated into the cells and then taken up by cells through endocytosis. In the dividing cells, the nuclear envelope is disappeared, and the DNA crystals can get direct entry into the nucleus of these dividing cells through this procedure. But in the non-dividing cells, DNA crystals cannot get direct entry into the nucleus, and they remain intact in the cytoplasm. So, this technique of DNA coprecipitation becomes limited while using non-diving cells.  

In the coprecipitation technique, the level of protein expression can easily be altered by changing the concentration of DNA that is being used to make DNA crystals. This technique has proved to be very effective in the experiments of live imaging that concentrate on a single cell. This method can also be utilized to find out the subcellular location of proteins and their colocalization with RNAs in different neural cells. 

Lipofection:

This method of transfection is based on gene delivery through positively charged lipid molecules. They interact with the nucleic acids that are negatively charged in nature and form liposomes. Often neutral helper lipids are combined with the positively charged lipid molecules to facilitate their fusion with the plasma membrane. A complex is formed between nucleic acids and lipid molecules. Later, this complex is internalized into the cell through the process of endocytosis. This method of transfection is effective in a variety of cell types involving primary neural cells and granule neurons. Moreover, the liposomes work in the serum-containing environment which results in lesser cytotoxic impacts of transfection. 

The transfection ability of lipofection varies from cell to cell but is remarkably high with cell lines. The Lipofection technique works better in the dividing cells, and its effectiveness is greatly reduced when applied to the non-dividing cells. To induce RNA interference, cytoplasmic transfer of small interfering RNAs via lipofection is enough. In non-dividing cells, various oligonucleotides, small interfering RNAs, and microRNAs can be delivered efficiently to the cytoplasm via lipofection. This technique minimizes the risks of causing genetic alterations and immune responses to other methods of transfection.  

Viral-based transfection methods:

Viruses have emerged as a rapid and efficient tool for delivering nucleic acids into the cells. They can efficiently transfect both dividing and non-dividing cells. They are considered the only viable option to transfect cells with nucleic acids. Different viruses contain different tropisms and this aids in the limitation of transgene expression into certain subsets of the cells As viruses are very diverse in nature, the selection of viral vectors greatly depends on the target cells, the gene of interest, and the purposes of the applied experiment. There is a list of viruses that are commonly used in the transfection technique: 

• Adenoviruses 
• Adeno-associated viruses 
• Herpes simplex viruses 
• Lentiviruses

Microinjection technique:

In this technique, glass capillaries are used to inject DNA or RNA into the cells. All those necessary materials that are not made in the cells can be injected into the cell through this technique. Microinjection can also target certain neural cells in mixed cell culture. This technique is limited because the direct entry of glass capillaries causes stress in the plasma membrane which minimizes the survival rate of certain neural cells.