Product Highlights:

  • A microscope magnifies objects, revealing details not visible to the naked eye. It can project the image to the eye or a camera for observation or recording.
  • Avantier’s diverse lineup, featuring achromatic, semi-apochromatic, and apochromatic options, ensures superior imaging quality. With magnifications ranging from 1X to 100X and numerical apertures up to 1.45, Avantier caters to a wide range of microscopy needs. 
  • Avantier excels in specialized objectives like polarizing, phase contrast, and metallurgical objectives.
  • Custom solutions by Avantier extend capabilities into UV, NUV, and NIR regions, to push the boundaries of precision microscopy for research, industry, and medical diagnostics.

 

The Complete Guide to Microscope Objective Lens

Table of Contents

What is a Microscope Objective Lens?

Microscope objective lenses, vital optical elements in microscopy, enable precise observation of specimens. Objective lens manufacturers offer a wide range of objective designs for specific needs: high power for detailed observation, scanning for broader views, oil immersion for high-resolution imaging, and long working distance for manipulation without compromising quality. Those objectives are designed with advanced construction techniques for high performance objectives with a spring loaded retractable nose cone assembly that protects the front lens elements and the specimen from collision damage.

Adding to these features, long working distance objectives allow ample space between the lens and the specimen, facilitating the manipulation of samples without compromising image quality. Infinity correction objectives utilize infinity-corrected optical systems, providing flexibility and compatibility with various microscopy accessories.

Numerical aperture, magnification, optical tube length, degree of aberration correction, and other important characteristics are typically imprinted or engraved on the external portion of the barrel for easy reference. These specifications help researchers select the appropriate objective for their experiments, ensuring optimal performance and total magnification when combined with the ocular lens. Specifications like numerical aperture and magnification are typically labeled on the barrel for easy reference. These lenses are indispensable in scientific research providing high powered optics essential for research.

In the following content, we delve intensively into the various components and features of microscope objective lenses, exploring their construction, functionality, and specialized designs that enable researchers to gain deeper insights into the microscopic world.

Components of a microscope

A microscope is an optical device designed to magnify the image of an object, enabling details indiscernible to the human eye to be differentiated. A microscope may project the image onto the human eye or onto a camera or video device. 

Microscopes are usually complex assemblies that include an array of lenses, filters, polarizers, and beamsplitters. Illumination is arranged to provide enough light for a clear image, and sensors are used to ‘see’ the object.

Although today’s microscopes are usually far more powerful than the microscopes used historically, they are used for much the same purpose: viewing objects that would otherwise be indiscernible to the human eye.  Here we’ll start with a basic compound microscope and go on to explore the components and function of larger more complex microscopes. We’ll also take an in-depth look at one of the key parts of a microscope, the objective lens.

 

In many microscopes, backlight illumination is favored over traditional direct light illumination due to the latter’s tendency to over-saturate the object under inspection. One specific backlight illumination technique employed in microscopy is Koehler illumination. This method involves flooding the object with light from behind using incident light from a source like a light bulb (see Figure 2). Koehler illumination utilizes two convex lenses, the collector lens and the condenser lens(or called field lens) , to ensure even and bright illumination on both the object and image planes. This design prevents imaging the light bulb filament, a common issue with direct light illumination. Backlight illumination is also commonly referred to as brightfield illumination.

Figure 1 the complete guide to Microscope Objective Lens
Figure 1

For brightfield illumination to be effective, there needs to be a variation in opacity across the object. Without this variation, the illumination creates a dark blur around the object, resulting in an image with relative contrast between the object’s parts and the light source. Typically, brightfield illumination allows clear visualization of each part of the object unless it is extremely transparent. In cases where transparency hinders feature distinction, darkfield illumination becomes useful.

 

Darkfield illumination directs light rays obliquely onto the object, avoiding direct entry into the objective. Despite this oblique angle, the rays still illuminate the object plane. The resulting darkfield illumination image achieves high contrast between the transparent object and the light source. In a darkfield setup, a light source forms an inverted cone of light that blocks central rays but allows oblique rays to illuminate the object (see Figure 3). This design effectively forces light to illuminate the object without entering the optical system, making darkfield illumination particularly suitable for transparent objects. In contrast, no rays are blocked in a brightfield illumination setup.

the complete guide to microscope objective lens
Figure 2
the complete guide to Microscope Objective Lens
Figure 3

Epi-illumination, a third form of illumination employed in microscopy, generates light from above the objective. This setup replaces the need for a Koehler illumination configuration, as both the objective and the epi-illumination source contribute to the illumination process. The compact structure  of epi-illumination is a significant advantage, as the objective serves as a primary source for a considerable portion of the illumination. Figure 4 provides a depiction of a frequently used epi-illumination setup, particularly common in fluorescence applications.

the complete guide to Microscope Objective Lens
Figure 4

Compound Microscope

While a magnifying glass consists of just one lens element and can magnify any element placed within its focal length, a compound lens, by definition, contains multiple lens elements. A relay lens system is used to convey the image of the object to the eye or, in some cases, to camera and video sensors.

A basic compound microscope could consist of just two elements acting in relay, the objective and the eyepiece. The objective relays a real image to the eyepiece, while magnifying that image anywhere from 4-100x.  The eyepiece magnifies the real image received typically by another 10x, and conveys a virtual image to the sensor. 

There are two major specifications for a microscope: the magnification power and the resolution. The magnification tells us how much larger the image is made to appear. The resolution tells us how far away two points must be to be distinguishable. The smaller the resolution, the larger the resolving power of the microscope. The highest resolution you can get with a light microscope is 0.2 um, but this depends on the quality of both the objective and eyepiece.

Both the objective lens and the eyepiece also contribute to the overall magnification of the system. If an objective lens magnifies the object by 10x and the eyepiece by 2x, the microscope will magnify the object by 20 times. If the microscope lens magnifies the object by 10x and the eyepiece by 10x, the microscope will magnify the object by 100x. This multiplicative relationship is the key to the power of microscopes, and the prime reason they perform so much better than simply magnifying glasses. 

In modern microscopes, neither the eyepiece nor the microscope objective is a simple lens. Instead, a combination of carefully chosen optical components work together to create a high quality magnified image. A basic compound microscope can magnify up to about 1000x. If you need higher magnification, you may wish to use an electron microscope, which can magnify up to a million times.

  • Eyepiece 
    • In the initial stages of microscope development, eyepieces are integral to the design as they provide the sole method for visually observing the object under examination. Presently, analog or digital cameras have taken on this role, projecting the object’s image onto a monitor or screen. Microscope eyepieces typically comprise a field lens and an eye lens, with various designs available, each capable of producing a broader field of view (FOV) compared to a single-element design.
  • Microscope Illumination 
    • Most microscopes rely on background illumination such as daylight or a lightbulb rather than a dedicated light source. In brightfield illumination (also known as Koehler illumination), two convex lenses, a collector lens and a condenser lens,  are placed so as to saturate the specimen with external light admitted into the microscope from behind. This provides a bright, even, steady light throughout the system
  • Objectives: Refractive
    • Refractive objectives have their name because the elements bend or refract light as it passes through the system. They are well suited to machine vision applications, as they can provide high resolution imaging of very small objects or ultra fine details. Each element within a refractive element is typically coated with an anti-reflective coating. A basic achromatic objective is a refractive objective that consists of just an achromatic lens and a meniscus lens, mounted within appropriate housing. The design is meant to limit the effects of chromatic and spherical aberration  as they bring two wavelengths of light to focus in the same plane. Plan Apochromat objectives can be much more complex, with up to fifteen elements. They can be quite expensive, as would be expected due to their complexity.
  • Objectives: Reflective 
    • A reflective objective works by reflecting light rather than bending it. Primary and secondary mirror systems both magnify and relay the image of the object being observed. While reflective objectives are not as widely used as refractive objectives, they offer many benefits. They can work deeper in the UV or IR spectral regions, and they are not plagued with the same aberrations as refractive objectives. As a result, they tend to offer better resolving power.

Key Concepts and Specifications

The majority of microscope objective specifications are conveniently displayed on the objective’s body, including information such as the objective design/standard, magnification, numerical aperture, working distance, lens to image distance, and cover slip thickness correction. Refer to Figure 5 for guidance on interpreting microscope objective specifications. This direct placement of specifications on the objective facilitates a clear understanding of its characteristics, a crucial aspect when integrating multiple objectives into an application. Any additional specifications, like focal length, field of view (FOV), and design wavelength, can be readily calculated or obtained from the vendor or manufacturer’s provided specifications.

Figure 5
  • Aberration Correction or Resolution: Spherical and chromatic aberrations limit the resolution of conventional microscopes. Lenses with a high degree of aberration correction result in high-resolution images over the entire field of view.
  • Conjugate Distance: Objectives are corrected for a specific projection distance. In finite conjugate optical design, light from a non-infinite source is focused down to a point. In infinity-corrected optical systems, light emitted from the specimen passes through the objective lens and enters the tube lens as an infinity parallel beam, forming a real image.
  • Numerical Aperture (NA): NA is the measure of its capability to gather light and to resolve fine specimen details at a fixed object. A lens with a high NA collects more light and can resolve finer specimen details at a fixed distance. These are the elements that determine resolution, depth of focus, and image brightness. The larger the numerical aperture, the higher the resolution and the brighter the image can be observed. The higher the magnification of the objective lens, the larger the numerical aperture.

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  • Magnification: A microscope’s ability to produce larger images is referred to as magnification. High magnification objectives provide extremely detailed images of specimens. The term magnification is often confused with the term resolution, which describes the ability of an imaging system to show detail in the object that is being imaged. While high magnification without high resolution may result in very small microbes visible, it will not allow observers to distinguish between microbes or subcellular parts of a microbe. Avantier has spent years designing and manufacturing to satisfy both magnification and resolution requirements simultaneously. This is the magnification of the intermediate image (inverted real image) for the specimen. In addition to low magnification (4x to 10x), medium magnification (20x to 50x), high magnification (100x or more), extremely low magnification (2.5x) (below) etc.
  • Field of View(FOV): FOV refers to the portion of the object captured by a microscope system. The size of the FOV is dictated by the objective magnification. When employing an eyepiece-objective system, the FOV initially observed through the objective is enlarged by the eyepiece for visual examination. In a camera-objective system, this FOV is transmitted onto a rectangular camera sensor. Due to the sensor’s shape, it can only capture a segment of the complete circular FOV from the objective. In contrast, the human eye’s retina can capture a circular area, encompassing the entire FOV. Consequently, the FOV produced by a camera-microscope system tends to be slightly smaller than that of an eyepiece-microscope system.

 Field of View Formula

For more details, read this article.

  • Cover Glass: Objectives are usually corrected for a specific cover glass thickness, with 0.17 millimeters being the standard. The thickness of the cover glass is numerically marked on the objective lens. There are three types: one for cover glass specimens, one for non-cover specimens, and one for both cover glass specimens and no cover glass specimens.
  • Immersion Medium: The main purpose of using different types of immersion medium is to minimize the refractive index between the objective and the sample. It is crucial to use the correct medium, such as water, oil, or air/dry, as specified by the objective.
  • Working Distance: The distance between the front end of the microscope objective and the surface of the specimen at which the sharpest focus is achieved. Proper positioning is important to obtain a good image at the specified magnification. The distance from the tip of the objective lens to the specimen surface when focused. The larger the numerical aperture of the objective lens, the shorter the working distance.
  • Parfocal Length: The distance from the mounting plane of the objective to the sample plane. 
  • Working Wavelength(s): Objectives are corrected for specific wavelengths, with shorter wavelengths yielding higher resolution.

Types of Objectives Lenses

Basic Classification and its Application

Type

Overview

Applications

Ach: Achromat

Corrected for chromatic aberration of two wavelengths (red and blue).

This is the most common method, but if you focus on the center of the field of view, the periphery becomes blurred, so it is not suitable for inspection photography.

  • General observation
  • Practice

Plan: Plan-Achromat

Corrected for wavelength chromatic aberration and field curvature aberration.

It is suitable for inspection photography because it focuses not only on the center of the field of view but also on the periphery, producing a flat image.

  • General observation
  • inspection
  • photograph

 

PlanFl: Plan Semi Apochromat

The chromatic aberration of the three wavelengths, with a slight chromatic aberration remaining in the purple, and the curvature of the field have been corrected. Also called fluorite.

In terms of performance, it is positioned between the plan achromat objective lens and the plan apochromat objective lens. High Grade type.

  • inspection
  • research

PlanApo: Plan-Apochromat

Chromatic aberration of three wavelengths and field curvature aberration are corrected.

It has a large numerical aperture, excellent resolution, and can obtain ideal images. Highest grade type.

  • inspection
  • research
Plan-Achromat Microscope Objective Lens
Flat-field-super-achromatic-objective
Achromat Microscope Objective Lens
Apochromats-Lenses
Plan-Apochromatic Microscope Objective Lens
Plan Semi Apochromatic Microscope Objective Lens
Plan Semi Apochromatic Microscope Objective Lens
Aberration Correction Basis
Aberration Correction Basis

Type

Overview

Achromat

Corrected for chromatic aberration of two colors (red and blue). This is the most common method, but if you focus on the center of the field of view, the periphery becomes blurred, so it is not suitable for photography.
Plan-Achromat Corrected for two-color (red and blue) chromatic aberration and field curvature aberration. It is suitable for photography because it focuses not only on the center of the field of view but also on the periphery, producing a flat image.

Fluorite

Corrected for chromatic aberration of three colors (red, blue, and violet) with minimal remaining aberration in the purple spectrum. This lens type offers improved color accuracy and resolution compared to achromats.

Plan Fluorite

Combines the correction of three-color chromatic aberration (red, blue, and violet) with field curvature aberration correction. This advanced lens design provides exceptional image quality across the entire field of view, making it ideal for precise microscopy applications requiring high resolution and accurate color rendition.

Plan Semi Apochromat

The chromatic aberration of the three colors (red, blue, and violet), with a slight chromatic aberration remaining in the purple, and the curvature of field have been corrected. Also called fluorite. In terms of performance, it is positioned between the plan achromat objective lens and the plan apochromat objective lens. High Grade type.
System Basis

Type

Overview

Reflective Objective Lenses

Microscope objective lenses that include two or more mirrors are called this type of objective that typically boasts a long working distance and zero chromatic aberration, and is useful for FTIR spectroscopy applications.
Refractive Objective Lenses Microscope objective made of transparent materials like glass. They use refraction, not reflection, to focus light onto the specimen. These lenses consist of multiple lens elements that correct for optical aberrations and ensure high-quality imaging.
Immersion Medium Basis

Type

Overview

Oil-immersion Objective Lenses

Most lenses are designed to work with air between the objective and cover glass, a higher numerical aperture can sometimes be achieved with other substances such as special oil with a specific refractive index.
Water immersion Objectives Lenses Water is used as the immersion medium between the objective and cover glass. This allows for a higher numerical aperture, improving resolution and image quality in microscopy applications.

Air immersion Objective Lenses

Air is used as the medium between the objective and cover glass. They are commonly used in standard microscopy setups and provide good resolution and image clarity.

Glycerol immersion Objective lenses

Combines the correction of three-color chromatic aberration (red, blue, and violet) with field curvature aberration correction. This advanced lens design provides exceptional image quality across the entire field of view, making it ideal for precise microscopy applications requiring high resolution and accurate color rendition.

Offerings by Avantier

Avantier’s Line-ups

Type of Objectives

Magnification

Numerical Aperture

Working Distance (mm)

Compatible Brands

Finite Correction Objectives

Achromatic, Semi-Plan Achromatic, Plan Achromatic1X, 4X, 10X, 20X, 40X, 60X, 100X0.1 – 1.250.13 – 15.8

Infinitely-Corrected Objectives
2.1 Tube Lens FL = 150mm

Semi-Plan Achromatic4X, 10X, 20X, 40X, 60X, 100X0.1 – 1.250.13- 5.02

Cover Glass Thickness Corrected
2.2 Tube Lens FL = 180mm

Semi-Plan Achromatic, Plan Achromatic. Inverted Micro, Fluorescent Objective for Laboratory Micro, Plan Semi Apochromatic, Semi-APO Fluorescent2X, 4X, 5X, 10X, 20X, 40X, 60X, 100X0.1 – 1.250.12 – 28

Cover Glass Thickness Corrected
2.3 Tube Lens FL=200mm

 

Plan, Apochromatic, Semi-APO Fluorescent, Semi-APO Metallurgical, Semi-Apochromatic, Plan Achromatic2X, 4X, 10X, 20X, 40X, 50X, 60X, 100X0.1 – 1.450.13 – 20.6

Compatible Tube Lens EFL:200mm

Plan Fluor EPI & EPI BD5X, 10X, 20X, 50X, 100X0.14 – 0.91 – 20Olympus equivalent

Polarizing Objectives

Polarizing4X, 10X, 20X, 40X, 100X0.1 – 1.250.2 – 30

Phase Contrast Objectives

Plan Phase Contrast, Semi-Apochromatic4X, 10X, 20X, 40X, 60X, 100X0.2 – 1.250.2 – 17.2

Semi-APO Metallurgical Objectives

Semi-APO Metallurgical5X, 10X, 20X, 50X, 100X0.15 – 0.91 – 20

Flat Field Achromatics Objectives

Infinitely Conjugated Ultra-Long Working Distance Flat Field Achromatic1X, 2X, 3.5X, 5X, 7.5X, 10X, 15X, 20X, 50X0.025 – 0.5513 – 41

Plan ApochromaticObjectives

Infinitely Far Conjugated Flat Field Achromatic, M Plan Apo NIR, LCD Plan Apo NIR. M Plan Apo NUV (HR)5X, 10X, 20X, 50X, 100X0.21 – 0.91.4 – 20Mitutoyo equivalen

Infinitely Conjugated Flat Field Apochromatic Near-Ultraviolet Objective

M Plan Apo NUV20X, 50X,0.42 – 0.6510 – 17 

Infinite Conjugated Ultra-Long Working Distance Bright Field Objective

M Plan Apo NIR, i Plan Apo HR, M Plan Apo HR NIR, M Plan Apo SL20X, 50X, 100X0.42 – 0.754 – 30Mitutoyo equivalent

Small Objectives (Lens diameter <4mm)

 3X, 7X0.45 – 0.70.01 – 1 
LCD Plan Apo NUV  

Microscope Objective Selection Guide

 

Microscope objectives are pivotal components in optical microscopy, especially in influencing image quality and resolution. Selecting the right objective is crucial for achieving optimal results in your microscopy applications. To guide you through the selection process, consider the following factors:

1. Working Distance
  • Working distance (WD) is the distance from the objective to the coverglass.
  •  Inversely proportional to numerical aperture (NA); higher NA often means lower working distance.
  • Consider long-working distance objectives for specific applications.
  • Resolution is determined by NA and illumination wavelength.
  • Higher NA provides finer resolution.
  • Choose NA carefully based on your application’s resolution needs.
  • Evaluate field number (FN) for the diameter of the field of view.
  • Modern objectives have FNs between 22 and 26.5mm.
  • Consider depth of field, which varies with numerical aperture.
  • Determine the size of your specimen.
  • Our microscope objectives offer a magnification range from 1.25x to 150x.
  • Consider the combined magnification with eyepieces for overall magnification.
  • Assess the numerical aperture (NA) of objectives.
  • Higher NA enables to gather more light, enhancing resolution and brightness.
  • NA ranges from 0.04 to 1.7; choose based on your specimen’s fine structures.
  • High NA objectives are recommended for weak fluorescence signals.
  • Avantier offers a range of objectives for fluorescence excitation across UV to NIR.
  • Choose objectives based on chromatic correction needs.
  • Achromat, semi-apochromat, and apochromat objectives for multichannel fluorescence.
  • Extended apochromats recommended for multichannel applications.
  • Consider observation methods beyond brightfield.
  • Dedicated objectives for darkfield, DIC, phase contrast, and polarization.
  • Select objectives based on the immersion medium: air, water, oil, or silicone.
  • Immersion mediums can enhance resolution; choose accordingly.
  • Dedicated objectives for advanced systems like confocal microscopy.
  • Choose objectives tailored to confocal, spinning disk confocal, multi-photon excitation, and TIRF microscopy.
  • Avantier offers a diverse range of objectives with varying aberration corrections.
  • Consider the crucial conjugate distance – finite for simple systems, and infinite for research-grade applications.

Choosing the right microscope objective is pivotal for optimal imaging performance. Consider your specific application requirements, utilize the provided guide, and explore Avantier’s diverse objective offerings to ensure accurate and reliable results in your microscopy endeavors.

Custom Microscope Objectives Solutions 

Avantier is a premier manufacturer of high performance microscope objective lenses, and we produce a wide range of quality microscope objectives for applications ranging from research to industry to forensics and medical diagnostics. We carry many types of objectives in stock, including apochromat objectives, achromatic objectives,  and semi apochromat objectives.  We can also produce custom objectives designed to work as desired in your target spectral range.

For custom microscope objective lenses, visit our microscope objective lenses page. 

If you’re interested in acquiring in-stock microscope objective lenses, please visit our ‘Stock – Microscope Objective‘ page.

Appendix: Advanced Microscopy Applications

Fluorescence Microscopy

Fluorescence microscopy is a powerful imaging technique used primarily in biomedical research to visualize and study samples labeled with fluorescent dyes or proteins at the microscopic level. The method relies on the phenomenon of fluorescence, where materials absorb light at a specific wavelength (excitation light) and then emit light at a longer wavelength (emission wavelength). A focused light source, such as a laser, is used to selectively excite fluorescent molecules within the sample. The emitted fluorescence is captured to form detailed images, providing valuable information about the sample’s internal structure and composition.

Confocal Microscopy

Confocal microscopy offers the capability to capture sharp images from a slender slice of a dense sample, minimizing background noise and reducing out-of-focus disturbances. Optical sectioning, widely employed in biomedical science and materials science, involves placing a sample on the microscope stage. An image is initially acquired at the primary focal plane, and subsequently, the stage or objective is adjusted vertically to capture images at successive focal planes.

the complete guide to microscope objective lens

IR Microscope

Infrared microscopy, alternatively referred to as infrared microspectroscopy, is a form of light microscopy that employs a light source transmitting infrared wavelengths to observe a sample’s image. In contrast to conventional optical microscopes utilizing absorbent glass optics, an infrared microscope incorporates reflective optics, enabling it to encompass the complete spectral range of infrared light.

Laser Ablation

Lasers find widespread applications, commonly employed to either (1) heat material onto a base or (2) ablate material off of a base. Laser ablation systems necessitate the integration of microscope components due to the precise manipulation of the laser beam, including focusing, bending, and reducing scattering. Typically, a laser ablation setup incorporates custom optics instead of off-the-shelf components, with the laser intricately designed into the system, as illustrated in Figure 14. The laser is strategically oriented in an epi-illumination design to leverage the microscope objective’s capacity to focus light at the object plane, generating exceptionally small spot sizes with minimal aberrations. Additionally, an eyepiece enables the user to visually locate the laser and ensure proper functionality. Filters are indispensable in shielding the user’s eyes from potential laser damage. Laser ablation setups, known for their superior precision compared to traditional surgical methods, find applications in medical and biological contexts.

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