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Figure 3. Use of immersion media matched to the objective can minimize the refractive index differences between the objective and the sample.

Each objective is designed for a specific immersion medium, which is marked on the objective. The main types of immersion media are air, oil, and water. It is important that you never put air objectives in oil or other liquids. Doing this will make the person in charge of the microscope really angry! The main purpose of using different types of immersion media is to minimize the refractive index differences that are present in the space between the objective and the sample. This includes the substrate (i.e., glass coverslip) that the sample is on and the imaging medium (i.e., buffer) that the sample is in. Minimizing this difference will result in better image resolution.

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Learn the correct magnification for your experiment and how to tell if your objective can be used with air, oil, or other immersion media.

Numerical aperture is a property of the objective that indicates how good the resolution can be in the image you collect (basically how much fine detail you can see).

Light will travel through different types of materials at different rates. When light travels through one material (such as air) and into another (such as water), the light is refracted. It appears bent. For instance, when you put a pencil in a glass of water and view the glass from the side, the pencil will look bent. This is because air has a different refractive index than water.

A microscope objective is composed of a complex set of lenses and optics, and different objectives are designed for different imaging tasks. Capturing good images relies on choosing the correct objective.

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“Small to midsize manufacturing entities in Vermont need access to VMEC’s kind of expertise because it is more difficult to cross-pollinate our work forces here with individuals that can bring in Best Industrial Practices”, states John Thrailkill, Ai Co-Founder and CEO. He adds, “Other regions of the country that have a more established manufacturing base have a much easier time of it. Keep up the good work!”

Magnification tells you the optical magnification the objective provides. The magnification you choose depends on what you want to see. The usefulness of magnification will be limited by your resolution. Making a big fuzzy blob of light even bigger won’t give you a better picture. Provided you have similar resolution at different magnifications, using higher magnification will allow you to see smaller things (such as organelles inside a cell) better. On the other hand, using a lower magnification will give you a better image of the big picture—such as a field of cells or interactions between cells.

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The objective is an essential part of the microscope and can greatly influence image quality. Objectives come with lots of information written on them, and most of it is written in code. But don’t worry; it’s easy to decipher.

Figure 2. Same field of cells captured at different magnifications. Each magnification can offer different information, and the best choice for your experiment will vary depending on what you want to know.

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Advanced Illumination (Ai) is an industrial lighting company, primarily serving the Machine Vision Industry. Founded in the Boston area in 1993, Ai was the world’s first LED lighting company. A developer and manufacturer of structured lighting and control electronics, Ai supports a wide variety of manufacturing activities around the globe. After relocating to Rochester, Vermont, in 1996, Ai employs 35 people full time.

Figure 4. The pencil appears bent or broken because the refractive indexes of water and glass are different than that of air.

Lots of times, you will hear people talk about the “NA” of an objective. “NA” stands for numerical aperture and its value partly depends on the refractive index of the material that is between the objective and the glass coverslip that your sample is on. In general, objectives with higher NA give you better resolution. Higher NA objectives often have higher magnification and use some sort of immersion medium. Immersion medium is used to alter the refractive index of the space between the objective and glass coverslip so that it is closer to the refractive index of the glass coverslip itself. This minimizes refraction and loss of light, ultimately giving you a better image.

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Ai’s sales had remained flat for six years while the company lost market share to a growing number of competitors. They were seeking to grow the business by doubling their sales over the next five years as well as changing their sales organization from an external to internal team. This would allow for improvements to their sales and marketing efforts where the sales leadership would have the opportunity to develop strategies to drive the organization forward. Realizing they needed the necessary help to redefine the sales organization and having worked with them on projects in the past, VMEC was contacted.

The Immersion medium is what's between the objective and the coverslip (or the bottom of the dish or flask that holds your sample).

Results of this strategic planning effort included increased sales of $600,000, cost savings of $400,000 and information systems investment of $100,000.

The working distance is the distance between the objective and the cover glass, or between the objective and the top (or bottom) of whatever vessel you are imaging through, when your sample is in focus. When you are imaging through something thin, like a cover glass, you can use objectives with shorter working distances. But when you are imaging samples that are in a thicker vessel, such as a plastic plate or dish, you will probably need an objective that has a longer working distance. The working distance of an objective is often written on the objective. The working distance of the objective in this example is 7.4 mm. It is considered to have an ‘extra-long working distance’ and is abbreviated as ELWD on the objective.

VMEC worked with Ai on initial planning to move the company’s sales effort from outside to inside, driving organizational changes and business transformations. Information gathered to support the company’s efforts included inputs from CoreValue intake surveys, market knowledge and customer feedback. Gathering and assigning input components as well as a strategic planning session with Ai followed. The latter was crucial so the newly formed team could work together to understand the big picture by stepping back and seeing their process as a system and ultimately drive performance.