Transmission Electron Microscopes typically use high energy electron beams transmitted through very thin samples in order to analyze the microstructure of materials. Electrons are accelerated at several hundred KV, resulting in wavelengths much smaller than that of light, and are focused with electromagnetic lenses for imaging. Images of samples generated are recorded on digital camera. The TEM facility at IIT-Delhi consists of a 200 KV TEM, with a high brightness field-emission gun (FEG) source whichproduces improved sensitivity and resolution compared to more traditional thermionic sources like LaB6 or Tungsten filaments. Collected images provide detailed information about the size, shape and morphological details of materials. This facility can also be utilized for high resolution analysis of the structure and organization of large biological molecules such as protein complexes, viruses etc through cryoelectron microscopy. This technique involves freezing biological material quickly in vitreous ice and imaging at temperatures less than -180°C. Accessory equipment for plunge freezing samples, plasma cleaning grids and holders are available.
i) Microscope: FEI TecnaiTF20: The FEI Tecnai TF20 is a high resolution Transmission Electron Microscope (TEM) equipped with a FEG source. It has a maximum accelerating voltage of 200 KV, a +/-70 degrees tilted computer controlled stage, and is equipped with a 4K x 4K Eagle CCD Camera with a 4-port readout and 15μm pixel size.Images may be collected in low dose mode for cryo samples. The microscope is suitable for collecting data on macromolecules, cryoelectron microscopy or tomography of single particles as well as semi-thick frozen sections.
ii) Plunge freezing device:
Make : FEI and Model : Vitrobot Mark IV The Vitrobot (Mark IV) is an automated device forvitrification of aqueous samples for cryo-TEM. Accessories such as liquid ethane bath and ethane temperature controller for plunge-freezing of grids, and liquid nitrogen workstation for cryotransfer are available.
iii) Plasma Cleaner: Model 1020 plasma cleaner (Fischione Instruments Inc) The Model 1020 plasma cleaner from Fischione Instruments is capable of cleaning both TEM support grids and TEM specimen holders.
iv) Others:Cryoholder (Gatan), Turbo pumping station (Gatan), Smartset cold stage controller (Gatan) Single-tilt holder for regular samples and cryotransfer holder for cryo samples are available. The latter is supplied with temperature controller and dry pumping station.
v) Softwares: EPU software from FEI is available for automated data collection. Softwares for single particle reconstruction such as EMAN and SPIDER are freely available.
Samples
Required Samples
| Sample | Sample Conc. | Sample State | Pretreatment | Conditions for Cryo-EM | Sample Size |
| Human Rhinoviruses | 1.0-2.0 mg/ml | Dissolve in 50mM Tris buffer, pH~7.2 | Glow Discharge/Plasm a Cleaning 30 sec | T=25°C, H=80%, Vol.=4µl, B=0.7- 0.9s | 30 nm |
| Flock House Virus | 1.0-3.0 mg/ml | 50mM Hepes, 5mM Cacl2 pH~7.4 | Glow Discharge/Plasm a Cleaning60 sec | T=11°C, H=99%, Vol.=3µl, B=3s | 30 nm |
| POPC Liposomes | 2.5 mg/ml | Unbuffered | Glow Discharge/Plasm a Cleaning60 sec | T=30°C, H=99%, Vol.=4µl, B=0.8- 1.0s | 10-300 nm |
| Microtubules | 10 µM | 1mM EDTA, 1mM Mgcl2,80mM Pipes KOH, pH~6.8 | Glow Discharge/Plasm a Cleaning30 sec | Not Establised | ~50 micrometer |
| Ribosomes | 1.0-2.0 mg/ml | 10mM Tris-Hcl pH~7.5, 1mM Mgcl2 | Plasma Cleaning 30 sec | T=4°C, H=100%, Vol.=4µl, B=6s | 20nm |
| RNA Polymerase | Optimum | 10 mM potassium phosphate,pH 7.9,1O mM 2mercaptoetha nol, 0.1 M EDTA, and 10% glycerol | Plasma Cleaning 60 sec | T=18°C, H=90%, Vol.=4µl, B=2.5s | 400-500 kDa |
| Proteins | 0.5-1 mg/ml | 20mM Sodium phosphate, 20mM Nacl | Glow Discharge/Plasm a Cleaning30 sec | Not Established | 18.7 kDa Monomer |
| Antibody | 0.1-1 mg/ml | 100mM Citrate pH~3.0, 100mM Nacl | Glow Discharge/Plasm a Cleaning30 sec | T=30° | 6-10 nm Monomer |
| Material Science | Varies from sample to sample | Well dispersed in a solvent like ethanol or water | UltraSonication for about 30min. to 1 hr. as per sample | For observation under EM, mount sample amount 5- 10 uL on 3mm Cu grid then dry it overnight | 2µm-5nm |
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1. Registration process (One Time): New Registration for external users.User Verification: Upload photo of Front side and back side of valid Identity card issued by the organization (only .jpeg format, file Size less than 1 Mb, height less than 1000 pixels, width less than 800 pixels). Undertaking: Download the undertaking template ( Download Link ). Duly filled, signed and stamped form should be uploaded in .pdf format. Confirmation: An email will be sent to the user after verification of the documents.
2.Before planning to avail the Instrument facility of CRF IIT Delhi: user must visit https://crf.iitd.ac.in -> Facility -> select the desired facility from left side navigation list. Read carefully all the relevant information such as required sample type, testing charges and instrument location. Be prepared with details of the samples.
3.Log in and Click on Booking Appointment Tab: All the information relevant to testing must be filled in instrument booking form. In case user does not provide the required information the booking is liable to be rejected. User can upload the relevant existing data of their samples that might help in the measurements.
4.Sample submission & Payment Process: Once the application is submitted, the in-charge will verify the details and approve the request.External users must wait for the approval before making the payment and submitting the samples to concerned lab. After approval the payment should be made by the user. The payment transaction details need to be submitted online before appointment date time.
5.CRF Users can pay amount online using following details:
Beneficiary/Customer's Name: IRD ACCOUNT IITD
Bank Account Number: 10773572600
Bank Name: State Bank of India Branch Name: IIT Branch
Branch Code: 01077
NEFT IFSC CODE / RTGS: SBIN0001077
please write "CRF _Instrument Name" in the Remarks section while paying. Save the receipt file and upload in the crfbooking system online against your application.Payment can also made in form of demand draft in favour of " IRD ACCOUNT IITD " Payable at New Delhi.6.IIT Delhi
GST Number : 07AAATI0393L1ZI
PAN Number : AAATI0393L
1 Registration process (One Time): New Registration for internal users.
Fill in all the details in Personal Information, Supervisor Information and User Login Credentionals.
Once submitted, an email to supervisor is sent for activation of account. Once activated by supervisor, you can do the instrument bookings.
2 Before planning to avail the Instrument facility of CRF IIT Delhi: user must visit https://crf.iitd.ac.in > Facilities > select the desired facility from left side navigation list. Read ca 
refully all the relevant information such as required sample type, testing charges and instrument location. Be prepared with details of the samples.
3. Log in on CRF Booking System and Click on Booking Appointment Tab: All the information relevant to testing must be filled in instrument booking form. In case user does not provide the required information the booking is liable to be rejected. User can upload the relevant existing data of their samples that might help in the measurements.
4 Sample submission & Payment Process: Once the application is submitted, the in-charge will verify the details and approve the request.
