A team led by AmitDutt from the Mumbai-based Tata Memorial Centre has turned to Raman Spectroscopy to detect RNA viruses present in saliva samples.
What is Raman Spectroscopy?
• Raman spectroscopy is an analytical technique where scattered light is used to measure the vibrational energy modes of a sample.
• It is named after the Indian physicist C. V. Raman who, together with his research partner K. S. Krishnan, was the first to observe Raman scattering in 1928.
• Raman spectroscopy can provide both chemical and structural information, as well as the identification of substances through their characteristic Raman ‘fingerprint’.
• Raman spectroscopy extracts this information through the detection of Raman scattering from the sample.
What is Raman Scattering?
When light is scattered by molecule, the oscillating electromagnetic field of a photon induces a polarisation of the molecular electron cloud which leaves the molecule in a higher energy state with the energy of the photon transferred to the molecule.
This can be considered as the formation of a very short-lived complex between the photon and molecule which is commonly called the virtual state of the molecule.
The virtual state is not stable and the photon is re-emitted almost immediately, as scattered light.
In the vast majority of scattering events, the energy of the molecule is unchanged after its interaction with the photon; and the energy, and therefore the wavelength, of the scattered photon is equal to that of the incident photon. This is called elastic (energy of scattering particle is conserved) or Rayleigh scattering and is the dominant process.
In a much rarer event Raman scattering occurs, which is an inelastic scattering process with a transfer of energy between the molecule and scattered photon.
If the molecule gains energy from the photon during the scattering (excited to a higher vibrational level) then the scattered photon loses energy and its wavelength increases which is called Stokes Raman scattering (after G. G. Stokes).
Inversely, if the molecule loses energy by relaxing to a lower vibrational level the scattered photon gains the corresponding energy and its wavelength decreases; which is called Anti-Stokes Raman scattering.
Quantum mechanically Stokes and Anti-Stokes are equally likely processes. However, with an ensemble of molecules, the majority of molecules will be in the ground vibrational level (Boltzmann distribution) and Stokes scatter is the statistically more probable process.
As a result, the Stokes Raman scatter is always more intense than the anti-Stokes and for this reason, it is nearly always the Stokes Raman scatter that is measured in Raman spectroscopy.
It has been reported that novel coronavirus is found in sufficient numbers in human saliva.
For the study, the researchers spiked saliva samples with non-infectious RNA viruses and analysed it with Raman Spectroscopy. They analysed the raw Raman Spectroscopy data and compared the signals with both viral positive and negative samples.
Statistical analysis of all the 1,400 spectra obtained for each sample, showed a set of 65 Raman spectral features was adequate to identify the viral positive signal.
Interestingly, most of the spectra were specific for the RNA molecule.
To minimise variability and automate the analysis of the Raman spectra for RNA viruses, they developed an automated tool — RNA Virus Detector — using a graphical user interface.
The tool can be used for detecting RNA virus from an individual or a group of samples in an unambiguous and reproducible manner, and is freely downloadable.
This tool, the first of its kind, takes raw data from a Raman Spectrometer analysis based on the 65-spectra signature and provides an objective output if viral RNA is present or absent in the sample.
This conceptual framework to detect RNA viruses in saliva could form the basis for field application of Raman Spectroscopy in managing viral outbreaks, such as the ongoing COVID-19 pandemic.
Since the tool can only identify RNA viruses and not identify the specific one, it can be used only for screening.
The RNA virus detected could be a common cold virus as well or any other RNA virus such as HIV. It doesn't look for COVID-19 viral-specific signature.
The advantage is that the tool can be taken to the field and people who test positive for RNA virus can be quarantined while another sample may be sent for validation using RT-PCR.
This whole process of data acquisition and analysis can be performed within a minute.
Since no additional reagent is needed there is no recurring cost. A portable (benchtop or handheld) Raman spectrophotometer installed at the port of entry such as airports or any point of care (in the field) can quickly screen passengers within minutes.