Hexamethyl Disilazane as an Alternative to Critical Point Drying

Critical point drying (CPD) is an established method of dehydrating biological tissue prior to examination in the Scanning Electron Microscope (SEM). This technique was first introduced to preserve three-dimensional structure of biological specimens for transmission electron microscopy, later it was used for obtaining dry specimens for SEM examination. Although CPD is a well-established method for drying specimens, there has been compelling evidence that it can damage soft specimens in two different ways: Structural integrity and Chemical Integrity. Over the years, use of the pharmaintermediate, Hexamethyl Disilazane (HMDS) in critical point drying is an established method of dehydrating biological tissue prior to examination in the SEM. Moreover, use HMDS in the imaging of cells on coal instead of CPD have led to negating the need for expensive equipment.

Studies on the applications of Hexamethyl Disilazane (HMDS) over Critical Point Drying (CPD):

• CPD and HMDS sample preparation techniques were used for testing the effects of both on the cervical cells on field emission scanning electron microscopy and energy dispersive X-ray. The results indicated that SEM imaging, elemental composition, and processing time for sample preparation with the HMDS technique were better than CPD technique for cervical cell preparation technique because in terms of weight percentages of carbon and oxygen element compositions in HMDS technique were higher than the CPD technique.

• CPD and HMDS samples used for preserving biological structures, revealed that samples prepared using CPD had tissue damage in the form of cracking. In addition there was excess amount of debris around the crack openings. Another potential result that placed HMDS as a superior drying agent is the time constraints that can be effective parameters while preparing more specimens; use of the Pharma Intermediate may be preferred over the traditional technique as it allows multiple samples to be dried simultaneously.

Preventing the gas/liquid interface is the main goal of critical point drying (as well as the alternative process of HMDS). Whenever a liquid evaporates into the gaseous phase, large surface tension phenomena occur. These surface tensions can harm fine surface details on the surface of the sample. CPD usually takes up to 3 hours to perform. Using Hexamethyldisilazane instead of CPD is quicker and can yield acceptable results on some samples. When the sample is dehydrated to 100% ethanol, a fifty/fifty mix of ethanol/HMDS can be placed on the sample followed by 2 or 3 exchanges of HMDS. After these exchanges, sample is in a fume hood where HMDS evaporates off. The vapor pressure of HMDS is such that surface damage is minimal.

The pharmaintermediate, Hexamethyl Disilazane is extensively used to replace the critical drying technique in sample preparation of tardigrades for SEM imaging because conventional procedures required CPD apparatus or machines to achieve the suitable temperature/pressure combination to completely dehydrate specimens, but with HMDS, the specimens can be dehydrated by simply bathing them in HMDS, ethanol solution, eliminating the use of any special equipment. All in all, the use of HMDS in SEM instead of CPD is preferred as it is safer, cheaper and more practical.

Hexamethyl Disilazane & Pharma Intermediate

Hexamethyl Disilazane, which is also known as HMDS, is a special organosilicon compound that is a derivative of ammonia. It is different from the traditionally known compound because it has trimethylsilyl groups in place of the two hydrogen atoms. It is a colorless liquid that is a popular reagent and precursor to many bases, making the compound a very popular ingredient in different types of organic synthesis and organometallic chemistry. HMDS has many uses as a chemical intermediate, release agent, lubricant, coupling agent, deactivator for chromatographic support materials, silylating agent in pharmaceuticals, adhesion promoter for photoresists on silicon (electronics), as reagent for temporary protection of reactive sites in manufacture of betalactum antibiotics, etc. Amongst these, the main applications of HMDS are in electron microscopy and in semiconductor electronics industry.

Major Applications of HMDS

HMDS represents an attractive to critical point drying (CPD) in the imaging of cells on coal, negating the need for expensive equipment. Coal, a popular choice in scanning electron microscopy, is easily fragmented into sub-micron particles, which can be problematic in critical point drying procedures.  Alternatively, HMDS was found to be easily used as an ideal substitute to critical point drying that takes place at the time of sample preparation. Even though critical point drying is a popular method of drying up biological specimens that are used for scanning electron microscopy, drying up of specimens by HMDS evaporation is also a very good alternative. Hexamethyldisilazane evaporation is not used frequently however experts have suggested that this is also a great method for drying up of specimens. Hardly any differences are observed during the process and furthermore this method is proved to be much more helpful as far as drying up of whole-mount cells for AFM and TEM are concerned.

Not only this, in the semiconductor industry, with the use of HMDS as semiconductor, companies have been able to enable powerful yet small chips inside computers, tablets and smartphones to work better on a consistent basis. HMDS is used to augment the adhesion of photoresist on silicon and SiO2 surfaces. The effectiveness of HMDS on adhesion is correlated with the reactivity of this compound with surface hydroxyl groups to form a new siloxane end product, i.e. Si-O-Si(CH3)3. This newly formed termination on the substrate renders the surface more hydrophobic in character and leads to greater wettability by photoresist. The latter condition is a crucial factor in good bonding. As a result of these altered characteristics due to the surface chemistry, the treated silicon surfaces become highly compatible with both negative and positive photoresists.

A few experiments were also made to verify the hydrophobicity and adsorption properties of HMDS thin films and the results pointed out that HMDS films can be used: for ultraviolet protection of flexible organic substrates, such as PP, for sensor and/or preconcentrator development, due to their adsorption properties, and in spatial applications due to resistance for O2 attack in hostile conditions, such as plasma etching. Thus, with many applications as described above, HMDS has found its way to be a major player as a Pharma Intermediate.