You are currently viewing 26 Difference Between Neutralization Assay and Hemagglutination Inhibition Assay

26 Difference Between Neutralization Assay and Hemagglutination Inhibition Assay

A neutralization assay is a laboratory procedure used to assess an antibody’s or antiviral agent’s capacity to reduce a virus’s or other infectious agent’s infectivity. In order to determine if the virus has been neutralized or inhibited from infecting target cells, the assay requires combining the virus with the test material (antibody or antiviral agent).

The ability of antibodies (typically against influenza viruses) to prevent the agglutination (clumping) of red blood cells by viral hemagglutinins is commonly assessed using the Hemagglutination Inhibition (HI) assay. Hemagglutinins, which are proteins on the virus’s surface, are in charge of the virus’s ability to attach to host cells.

An HI assay follows identical procedures to a general neutralization assay, but with a focus on preventing red blood cell agglutination. The test entails:

The virus is made, and serum from a subject who has been exposed to the virus or received a vaccination against it is obtained.

To produce a variety of concentrations, the serum is diluted in steps.

A predetermined virus concentration is combined with diluted serum. Antibodies to the virus will bind to the viral hemagglutinins and stop them from agglutinating red blood cells if the serum has these antibodies present.

 Red blood cells are introduced following incubation. Red blood cells will aggregate if the virus is successful in doing so, creating observable clumps.

The level of agglutination is monitored and noted in the agglutination readout. In comparison to controls without the serum, there will be reduced or no agglutination if the serum’s antibodies are successful in neutralizing the virus.

The hemagglutination inhibition titer is the highest dilution of the serum that inhibits hemagglutination. It gives an indication of the antibody’s capacity to prevent the virus from agglutinating red blood cells.

In virology and immunology, neutralization tests and HI assays are crucial techniques for determining the effectiveness of antibodies, vaccinations, and antiviral medicines. They support the development of treatments and preventive measures by assisting researchers in understanding how well these drugs work to stop viral infections.



Neutralization Assay

Hemagglutination Inhibition Assay



Measures the ability of antibodies to neutralize a virus or toxin

Measures the ability of antibodies to inhibit the agglutination of red blood cells



Determines the presence and potency of neutralizing antibodies

Measures the quantity of specific antibodies in a sample



Typically used for viral pathogens or toxins

Commonly used for detecting antibodies against certain antigens, including viruses



Involves assessing the reduction or prevention of viral infection

Focuses on the prevention of red blood cell agglutination


Virus or Antigen

Requires a live virus or toxin to perform the assay

Uses red blood cells as the antigen


Viral Growth

Requires a cell culture system for viral growth

Does not require viral growth or a cell culture system



Does not involve agglutination of cells

Focuses on inhibiting red blood cell agglutination



Determines if antibodies can block viral entry, replication, or other functions

Checks if antibodies can prevent red blood cell clumping



Typically, the readout is based on a reduction in virus-induced cytopathic effects

Readout is based on the absence of visible agglutination


Assay Duration

Can take several days to perform

Can be completed relatively quickly


Viral Titer Estimation

Not used for estimating viral titer

Can be used to estimate antibody titer


Antibody Detection

Not primarily used for detecting antibodies

Mainly used to detect antibodies in a sample


Viral Strain Specificity

Can be used to determine the specificity of neutralizing antibodies

May not provide information about antibody specificity


Suitable for All Viruses

May not work for all viruses due to the need for specific cell lines

Can be adapted for various viruses and antigens


Clinical Applications

Commonly used in vaccine development and serology testing

Primarily used in serology testing and blood typing



Generally highly sensitive to neutralizing antibodies

Sensitive to the presence of specific antibodies


Viral Load Estimation

Does not estimate viral load

Does not estimate viral load


Viral Variants Detection

Can be used to assess the efficacy against viral variants

Not typically used to detect viral variants


Requirement for Red Blood Cells

Doesn’t require the use of red blood cells

Depends on the presence of red blood cells


Viral Species Specificity

May be specific to certain viral species or strains

Not specific to viral species or strains


Vaccine Testing

Used to evaluate the efficacy of vaccines

Not used to evaluate vaccine efficacy


Antigen-Antibody Interaction

Assesses the blocking of virus-antibody interactions

Measures the inhibition of antigen-antibody reactions


Commonly Used in

Virology and immunology research

Blood banks and serology laboratories


Examples of Use

Used for assessing immunity to influenza, HIV, and other viral infections

Used for ABO and Rh blood typing and measles antibody detection


Risk of Contamination

Requires strict biosafety measures due to live virus use

Less risk of contamination as it doesn’t involve live viruses


Result Interpretation

Focuses on the reduction in viral infection or cytopathic effects

Concentrates on the prevention of red blood cell agglutination

Frequently Asked Questions (FAQ’S)

Q1. What purposes do neutralization assays serve?

Neutralization tests are used to assess the efficacy of vaccines, check the strength of antiviral medications, research the immune response to infections, and comprehend the neutralization properties of antibodies made in response to vaccination or naturally occurring illness.

Q2. What are the essential procedures for carrying out a neutralization assay?

Preparing viral samples, incubating the virus with the test material, introducing the mixture to target cells, enabling infection to take place, and calculating the decline in infection rates relative to control samples are the essential processes.

Q3. Neutralization tests can be used to research what kinds of viruses?

Numerous viruses, including influenza, HIV, hepatitis viruses, coronaviruses (such as SARS-CoV-2), and more can be studied using neutralization assays.

Q4. What role does the HI assay play?

The HI test is frequently used in influenza research and vaccine development to gauge the amount of antibodies that neutralize various viral strains. It aids in determining how well a vaccine works to produce defense-enhancing antibodies.

Q5. Do HI assays target influenza viruses specifically?

The HI assay can be modified for various hemagglutinating agents, even though influenza viruses are the ones most frequently associated with it. However, depending on the drug being examined, different methods and reagents may be used.

Q6. What procedures are followed when doing an HI assay?

Preparing red blood cells, dilution and incubation of test serum or antibodies with the virus, addition of red blood cells, and watching for agglutination are the main processes. The level of hemagglutination inhibition is determined by the maximum dilution that does not cause agglutination.

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Hi, I'm the Founder and Developer of the Serology Test, a blog truly devoted to Medics. I am a Medical Lab Tech, a Web Developer and Bibliophiliac. My greatest hobby is to teach and motivate other peoples to do whatever they wanna do in life.

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