10 Frequently Asked Questions About Elisa Kits

When carrying out an ELISA immunoassay, there are certain doubts or questions that are frequently repeated when carrying out our experiment.

In this week’s entry, we bring you a compilation of frequently asked questions about ELISA kits prepared by Biomatik , which can help you answer some questions.

Let’s start!


Although certain kits may have specific storage conditions that will be described in each case, in general, the components of the ELISA kits can be stored refrigerated (4ºC) or frozen (-20ºC).


The reagents must be prepared 10 minutes before use. When first used, reagents should be concentrated by centrifugation at the bottom of the tube.

Frequently, the amount of reagent in the tube is usually greater than the specified, so it is recommended to measure the amounts to be added accurately with a pipette or similar.


The well strips of the plates are mobile, so it is recommended to store those strips that are not going to be used immediately between 2-8ºC and in dark conditions.


All samples should be added over a 5 minute period. The time between each sample addition should be uniform to ensure consistency of the reaction.


Proper use of new, clean plate sealers must be ensured to avoid sample evaporation and contamination. When moving the plate, you must keep your hand steady so as not to spill the liquid. In order to maintain a constant temperature of 37ºC, excessive opening of the incubator door should be avoided.


The same volume of buffer should be used per well, ideally with a multichannel pipette. When using an ELISA plate washer, it must be ensured that it is clean and free from contamination. The plates should be gently tapped face down on the filter to dry them properly.


In order to avoid reading errors due to accumulation of precipitate, the plate should be read within 5 minutes of adding the stop solution. Make sure that the microplate reader is properly configured and the filter is properly calibrated. On the other hand, avoid mixing reagents with different lot numbers.


Out-of-standard curve results are not supported. Only those results that fall within the standard curve are reproducible and therefore accurate, according to the kit.


If the values ​​are above the standard curve, the samples should be diluted to fit the detection range of the kit.


You need to make sure that the target protein is expressed in your sample. In case the expression level is low, an attempt will be made to increase the amount of sample used. In some cases it may be necessary to resort to a higher sensitivity assay. It is also important to confirm that the positive control being used is within the detection range of the assay.

How To Reconstitute Lyophilized Peptides

The most stable way to store peptides for long periods of time is to keep them lyophilized, away from light and at low temperatures. However, when it comes time to use them, the work of reconstituting lyophilized peptides can be a headache.

Achieving the correct dissolution of the peptides is essential for the proper development of the assay, since otherwise peptide losses and / or inaccuracies may occur when calculating their concentration.

In this post we bring you some tips and guidelines that can help you reconstitute lyophilized peptides, and optimize their solubility .

Before proceeding to reconstitute lyophilized peptides, it is worth remembering some general considerations :

  • As a previous step to reconstitution, it is always convenient to centrifuge the vials containing the lyophilized peptide so that the complete sample remains in the pellet and no material is lost.
  • Before reconstitution, the peptide must be at room temperature .
  • It is recommended to test the solubility in a certain medium with a small amount of peptide , until confirming that it dissolves correctly.
  • The solvent of choice must be compatible with the experimental application and must not react or promote degradation of the peptide.
  • In general terms, the solubility of a peptide will be determined by its net charge , therefore:
    • Acid peptides can be resuspended in basic buffers.
    • The basic peptides will be resuspended in acidic solutions.
    • Hydrophobic and neutral peptides should be dissolved in small amounts of some organic solvent such as DMSO, methanol, acetic acid, isopropanol, etc.
  • In order to improve solubility, complementary techniques such as sonication can be used , if necessary.

When choosing the optimal solvent to reconstitute lyophilized peptides, it is necessary to take into account the two fundamental factors that will influence its solubility : the length of the peptide and its net charge.


As a general rule, peptides of less than 5-6 amino acids are usually soluble in aqueous solution. It should be remembered that distilled and sterile water should always be used.

For larger peptides, the solubility will be defined by their net charge and hydrophobicity, as detailed in the second point.


To calculate the net charge of the peptide, the following procedure will be followed, and the obtained values ​​will be added:

  • A value of -1 will be assigned for each acidic residue (Asp, Glu or C-terminal -COOH) that contains the peptide.
  • A value of +1 will be assigned for each basic residue (Arg, Lys, His or N-terminal -NH2) that contains the peptide.

Depending on the net charge value obtained, we will act as follows:

  • If the net charge of the peptide is negative (acid peptide):
    • First, attempts will be made to reconstitute the lyophilized peptide in water or PBS.
    • If that first option fails, NH4OH will be added (except in cases where the peptide contains Cys residues).
    • If the peptide still does not dissolve, an organic solvent such as DMSO will be added until it dissolves, and then it will be diluted to the desired concentration.
  • If the net charge of the peptide is positive (basic peptide):
    • First, an attempt will be made to reconstitute the lyophilized peptide in water.
    • If that first option fails, an attempt will be made to dissolve the peptide in a 25% acetic acid solution.
    • If the peptide still does not dissolve, an attempt will be made to solubilize in a small amount of DMSO and then dilute to the desired concentration.
  • If the net charge of the peptide is equal to 0 (neutral peptide):
    • First, attempts will be made to reconstitute the lyophilized peptide in an organic solvent.
    • In cases where the peptide is highly hydrophobic, an attempt will be made to solubilize in a small amount of DMSO and then it will be diluted with water to achieve the desired concentration.

Although in many cases when reconstituting lyophilized peptides these can be dissolved without problem in aqueous solutions, it is important to know the characteristics of the amino acids that compose them to be able to predict the solubility in each case. We hope that this entry has cleared some doubts about it.

Mobile applications addressing violence against women: a systematic review.

Violence against ladies is a urgent world well being drawback that’s being met with a new intervention strategy-mobile applications. With this systematic assessment, we offer an preliminary evaluation and useful categorisation of apps addressing violence against ladies.

We carried out a systematic on-line search conforming with Preferred Reporting Items for Systematic Reviews and Meta-Analyses tips to establish apps addressing violence against ladies in 5 World Bank areas (Europe and Central Asia; North America, Latin America and the Caribbean; Middle East and North Africa; South Asia; and sub-Saharan Africa). Applications with location of initiation in talked about areas and ≥100 downloads have been included.

Data on sector, goal group(s), 12 months of launch, location of initiation and implementation have been extracted. By technique of a structured qualitative content material evaluation, applications have been then categorised in accordance with their primary features.

ResultsOf 327 related applications, 171 have been included into the systematic assessment and assigned to one in every of 5 recognized classes of primary features, respectively: emergency, avoidance, schooling, reporting and proof constructing, and supporting apps. The largest proportion (46.78%) consisted of emergency apps, adopted by schooling, reporting and proof constructing, supporting and avoidance apps in descending order.

With regards to the geographical distribution of app classes, vital (χ2(20)=58.172; p=0.000) variations among the many included areas have been discovered.An enormous proportion of apps addressing violence against ladies primarily draw on one-time emergency or avoidance options, versus extra preventative approaches. Further analysis is important, critically contemplating questions of information safety, private security and efficacy of such cell well being interventions.

Mobile applications addressing violence against women: a systematic review.
Mobile applications addressing violence against ladies: a systematic assessment.

Cystic echinococcosis within the Eastern Mediterranean area: Neglected and prevailing!

Cystic echinococcosis (CE) is distributed worldwide, extending from China to the Middle East and from Mediterranean nations to the sub-Saharan Africa and South America. According to WHO, a million folks world wide are affected by CE with an estimated burden of 183,573 DALYs.

The annual financial burden of the illness resulting from therapy prices and CE-related livestock losses has been estimated at US$ three billion. CE is endemic in all nations throughout the WHO Eastern Mediterranean Regional Office (EMRO).

The area, which incorporates a lot of the Middle East and North Africa, is likely one of the most historic foci of the home cycle of CE and is acknowledged as one of many main hotspots of CE. There are 22 nations within the EMRO, the place about 688 million persons are residing susceptible to CE.

In many EMRO nations, little is thought about CE epidemiology and transmission. WHO included echinococcosis in a listing of 17 uncared for tropical illnesses (NTDs) and 12 uncared for zoonotic illnesses (NZDs).

Accordingly, completely different regional workplaces of WHO organized a number of initiatives for CE management and prevention. WHO’s Western Pacific regional workplace thought-about echinococcosis as one of many area’s main well being subjects, and a number of other preventive measures have been carried out within the American area with the help of Pan AmericaHealth Organization (PAHO) in Argentina, Peru, Uruguay, and Chile. Although CE is endemic in all 22 EMRO nations, surprisingly, CE is absent from the well being subjects listing of illnesses and situations on this area.

Therefore, CE clearly requires additional consideration within the WHO EMRO agenda, and the necessity for elaboration of particular measures for CE management is changing into obvious in EMRO nations, the place substantial collaborations among the many member states and WHO EMRO is of paramount significance.

Major subjects of collaborative actions embrace coaching packages and well being communication on completely different facets of CE management, evaluation of CE burden, nationwide and worldwide surveillance and illness registry programs, technical help to advertise epidemiological research for amassing baseline information, cost-benefit evaluation of management interventions, and intersectoral cooperation among the many agriculture, veterinary, medical, and well being sectors.

Creating an Age-Friendly Public Health System.

Background and ObjectivesThe public well being system in America-at all levels-has comparatively few specialised initiatives that prioritize the well being and well-being of older adults. And when public well being does tackle the wants of older adults, it’s usually as an afterthought.

In session with leaders in public well beingwell being care, and ageing, an revolutionary Framework for an Age-Friendly Public Health System (Framework) was developed outlining roles that public well being might fulfill, in collaboration with ageing providers, to handle the challenges and alternatives of an ageing society.

Research Design and MethodsWith management from Trust for America‘s Health and The John A. Hartford Foundation, the Florida Departments of Health and Elder Affairs are piloting the implementation of this Framework inside Florida’s county well being departments and on the state degree.

The county well being departments are increasing knowledge assortment efforts to determine older grownup wants, creating new alliances with ageing sector companions, coordinating with different businesses and neighborhood organizations to implement evidence-based packages and insurance policies that tackle precedence wants, and aligning efforts with the age-friendly communities and age-friendly well being methods actions.

Results and Discussion and ImplicationsThe county well being departments in Florida taking part within the pilot are leveraging the Framework to increase public well being follow, packages, and insurance policies that tackle well being providers and well being behaviors, social, and financial elements and environmental situations that enable older adults to age in place and reside well beingier and extra productive lives. The mannequin being piloted in Florida will be tailor-made to fulfill the distinctive wants of every neighborhood and their older grownup inhabitants.

Creating an Age-Friendly Public Health System.
Creating an Age-Friendly Public Health System.

Enfrentar las desigualdades en salud en América Latina: el rol de la protección social.

After greater than a decade of progress in numerous areas of social growth, since 2015 poverty has elevated, labor market indicators have deteriorated, and the discount of revenue inequality has stagnated in Latin America.

These developments are of concern as they’ll have an effect on well being indicators and exacerbate profound well being inequalities. This scenario calls for built-in coverage responses that may create synergies between completely different sectors.

There is rising recognition of the position of social safety within the eradication of poverty and the discount of inequality. Various social safety mechanisms buffer towards the prices of accessing well being providers straight and not directly.

By increasing protection and common entry, promotion and prevention actions in well being and vitamin, and essentially, the combat towards poverty, inequality, and exclusion, social safety performs a elementary position in guaranteeing the precise to well being and overcoming inequalities on this space.

The discount of inequalities in well being must be a precedence for all nations, and a manner ahead in that course is to advertise the development and strengthening of common social safety methods.

Violence against children in Latin America and Caribbean countries: a comprehensive review of national health sector efforts in prevention and response.

BACKGROUNDViolence against children (VAC) stays a world downside. The health sector has a chance and duty to be half of the multi-sector collaboration to forestall and reply to VAC.

This review aimed to evaluate the health sector’s response to VAC amongst Latin American & Caribbean (LAC) nations, notably because it pertains to bodily violence, sexual violence, and neglect.METHODSNational protocols for the identification and provision of health care to youngster survivors of violence, abuse and neglect have been solicited in partnership with UNICEF and PAHO/WHO nation workplaces inside the LAC area.

A parallel systematic review was undertaken in January 2015 to review research revealed in the final 10 years that describe the regional health sector response to VAC.

RESULTSWe obtained health sectors pointers/protocols associated to VAC from 22 of 43 (51 %) nations and reviewed 97 revealed articles/reviews that met the review inclusion standards. Country protocols have been offered in Spanish (n = 12), Portuguese (n = 1), and English (n = 9). Thematic areas of nation protocols included:

1) figuring out indicators and signs of VAC,

2) offering patient-centered care to the sufferer, and

3) fast remedy of accidents associated to VAC.

The systematic review revealed that health professionals are sometimes unaware of national protocols and lack coaching, sources, and assist to reply to instances of VAC. Further, there may be restricted coordination between health and social safety providers.CONCLUSIONSVAC stays a world, public health precedence.

Health professionals are well-positioned to establish, deal with and refer instances of VAC to applicable establishments and community-based companions. However, poor protocol dissemination and coaching, restricted infrastructure, and insufficient human sources problem adherence to VAC pointers.

Violence against children in Latin America and Caribbean countries: a comprehensive review of national health sector efforts in prevention and response.
Violence against children in Latin America and Caribbean nations: a comprehensive review of national health sector efforts in prevention and response.

Human sources: the Cinderella of health sector reform in Latin America.

Human sources are a very powerful property of any health system, and health workforce issues have for many years restricted the effectivity and high quality of Latin America health methods.

World Bank-led reforms geared toward rising fairness, effectivity, high quality of care and consumer satisfaction didn’t try and resolve the human sources issues that had been recognized in a number of health sector assessments. However, the 2 most necessary reform insurance policies – decentralization and privatization – have had a damaging influence on the circumstances of employment and prompted opposition from organized professionals and unions.

In a number of nations of the area, the workforce grew to become a very powerful impediment to profitable reform.This article is predicated on fieldwork and a review of the literature.

It discusses the explanations that led health staff to oppose reform; the institutional and authorized constraints to implementing reform as initially designed; the mismatch between the kinds of personnel wanted for reform and the supply of professionals; the deficiencies of the reform implementation course of; and the regulatory weaknesses of the area.

The dialogue presents workforce methods that the reforms may have included to realize the meant objectives, and the necessity to keep in mind the values and political realities of the nations. The authors recommend that autochthonous options usually tend to succeed than options imported from the skin.