Evaluating the reliability of the sickling test and peripheral blood film method for screening of sickle cell disease: a study protocol

Tejaswini Bawane; Sahitya Vodithala

doi:10.12688/f1000research.139432.1

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Study Protocol

Evaluating the reliability of the sickling test and peripheral blood film method for screening of sickle cell disease: a study protocol

[version 1; peer review: 1 approved with reservations]

Tejaswini Bawane ¹, Sahitya Vodithala³

PUBLISHED 11 Sep 2023

Author details Author details

¹ Pathology, Datta Meghe Institute of Higher Education and Research, Sawangi (M), Wardha India, Wardha, Maharashtra, 442001, India
² Pathology, Datta Megha Institute of Higher Education and Research Sawangi (M) Wardha. India, Wardha, Maharashtra, 442001, India
³ Pathology, Datta Meghe Institute of Higher Education and Research, Sawangi (M), Wardha. India, Wardha, Maharashtra, 442001, India

Tejaswini Bawane
Roles: Investigation, Methodology, Validation, Visualization, Writing – Original Draft Preparation

Sahitya Vodithala
Roles: Investigation, Methodology, Visualization

OPEN PEER REVIEW

REVIEWER STATUS

This article is included in the Datta Meghe Institute of Higher Education and Research collection.

Abstract

Background: Mutations in beta-globin are the cause of a widespread condition known as sickle cell disease (SCD). Sickle cells cause organ damage, hemolysis of blood cells, weakness, and sometimes even death. SCD can be effectively managed and mortality can be reduced through early detection. This protocol’s study aims to evaluate the sensitivity and specificity of the sickling test, peripheral blood smear, and hemoglobin electrophoresis for SCD screening.
Methods: In this study,75 cases of SCD will be selected. 5ml blood will be drawn into a dipotassium ethylenediaminetetraacetic acid tube for testing of SCD. Testing will involve: the sickling test, where an equal volume of K2EDTA blood will be mixed with 2% sodium metabisulfite; the peripheral blood smear, where Leishman’s stain will be placed on a thin film of blood for 2 minutes; and Hb electrophoresis, where equal parts of distilled water and packed cells will be combined and then the mixture will be centrifuged.
Results: We will be comparing between the sickling test and the peripheral blood film method, and we will conclude depending upon the results as to which test is better.
Conclusions: Using peripheral blood smear images, SCD can be quickly diagnosed. A peripheral smear-based differential diagnosis may be possible but requires special tests such as hemoglobin electrophoresis to confirm the diagnosis. Therefore, early diagnosis can help initiate transfusion therapy and create a better prognosis.

Keywords

Sickle cell disease (SCD), Anemia, Sickling test, Peripheral blood smear, Hemoglobin electrophoresis.

Corresponding author: Tejaswini Bawane

Competing interests: No competing interests were disclosed.

Grant information: The author(s) declared that no grants were involved in supporting this work.

Copyright: © 2023 Bawane T and Vodithala S. This is an open access article distributed under the terms of the Creative Commons Attribution License, which permits unrestricted use, distribution, and reproduction in any medium, provided the original work is properly cited.

How to cite: Bawane T and Vodithala S. Evaluating the reliability of the sickling test and peripheral blood film method for screening of sickle cell disease: a study protocol [version 1; peer review: 1 approved with reservations]. F1000Research 2023, 12:1124 (https://doi.org/10.12688/f1000research.139432.1) First published: 11 Sep 2023, 12:1124 (https://doi.org/10.12688/f1000research.139432.1) Latest published: 11 Sep 2023, 12:1124 (https://doi.org/10.12688/f1000research.139432.1)

Introduction

Red blood cells contain a pigment called hemoglobin, which carries oxygen to the tissue.¹ Defective hemoglobin structure is the cause of the hereditary blood disorder known as sickle cell disease (SCD).² In the year 1910 Herrick first described the sickle shaped red blood cell (RBC).³ A multisystem disease termed sickle cell anemia (SCA) is associated with acute illness and progressive organ damage.⁴

Point mutations cause SCA. In codon 6 of the HBB gene, which encodes a subunit, adenine is switched out for thymine (GAG > GTG). Nucleotide substitution has occurred. Hemoglobin S (Hgb S) is produced as a result of an amino acid change in which valine takes the place of glutamic acid. Sickle cells become rigid and less soluble when Hgb S polymerizes in anoxic conditions.⁴^,⁵ Hemolyticanemia is the result of this, which hastens cell death and reduces cell life by about 75%.⁶

Anaemia is not a diagnosis, but a finding that needs additional investigation. Anaemia is defined as a reduction in hemoglobin concentration below the lower limit of normal with regard to age and gender. In addition to low hemoglobin levels, a few other parameters should be taken as well when researching anemia. The initial investigations for these individuals begin with red cell indices, red cell count, and morphological evaluation of peripheral blood film.⁷

This disease is common in the tribal population of central India including Andhra and Telangana. The frequency of the sickle cell gene in the Indian population varies from 2-3.5 %.⁸

Though hemoglobin electrophoresis is the gold standard for diagnosis of SCD, peripheral smear examination remains an invaluable tool in preliminary diagnosis in unsuspected cases and those in sickle crisis.⁹ Early diagnosis, complete treatment, and general medical care have increased the life expectancy of patients with SCD in developed countries. Early detection, therefore, helps in effective disease management.¹⁰

Rationale

Sickle cell disease is a hemoglobinopathy caused by a point mutation in the ß globin gene that leads to the production of hemoglobin S, which polymerizes under deoxygenated conditions and causes red blood cells (RBCs) to form a sickle shape.¹¹^–¹³ Sickle RBCs result in hemolysis, vaso-occlusive pain crises, acute chest syndrome, post-operative infections, congestive heart failure, cerebrovascular accident and endothelial damage.¹⁴ Of all the screening tests used for screening cases of SCD, the sickling test is the most commonly used. However, these basic screening tests are plagued by a lot of disadvantages which may range from technical to human errors. The changes in the edge, location, shape, and size of the cells make blood film analysis extremely difficult. A computerized approach has been created as a result to provide a simple technique for identifying the type of anemia. The peripheral blood smear is an imperfect test, It is a formative hematological test that depends on the pathologist’s skills, and there aren’t many skilled pathologists to choose from. Haemoglobin electrophoresis, on the other hand, does not suffer from all these disadvantages and gives accurate results. Hemoglobin electrophoresis is the gold standard for the diagnosis of SCD. The routine protocol followed in the diagnosis of SCD is to do a screening test first and then go for a confirmatory test thus delaying the final diagnosis. The present study aims to employ both these tests for the screening cases of SCD and reliability with each other. Employing a superior diagnostic test will greatly reduce delays in diagnosing cases of SCD to a considerable extent.¹⁵

Recently, many studies have been done on techniques for the detection of SCD. Wjdan A. Arish et al.in (2021) studied the various prospective approaches that could be used to aid in the early identification of SCD, as well as the present and new methods for sickle cell disease detection.¹⁶

Emmanuel Gbenga Dada et al.’s (2022) study reported that traditional methods for diagnosing SCD are expensive, error-prone, time-consuming, and necessitate the help of medical professionals. The development of affordable and controllable methods for the early identification and diagnosis of SCD is thus required immediately.¹⁷

Protocol

Aim

To evaluate the reliability of the sickling test and peripheral blood film method for screening of sickle cell disease.

Objectives

1. To evaluate the sensitivity and specificity of the sickling test, peripheral blood smear, and hemoglobin electrophoresis for SCD screening.
2. To diagnose SCD on peripheral blood smear examination.
3. To screen for SCD of sickling.

Study design

This will be a descriptive, laboratory-based, observational and comparable study.

Ethical considerations

Ethical approval for the study was obtained from the institutional ethics committee of Datta Meghe Institute of Higher Education and Research (Deemed to be University) (ref. No: DMIHER (DU)/IEC/2023/24), date of approval: 21/07/2023. A written participant information sheet will be given regarding the details of the study, and it will be explained to participants before enrolment in the study. Their involvement benefits and harms will be explained to the participants. Written informed consent from the participants will be obtained before involving them in the study.

Study settings

The following study will be conducted in the Department of Pathology (Central Clinical Laboratory), AVB Rural Hospital, Sawangi Meghe (Wardha), India.

Study participants

Inclusion criteria

1. Patients aged 6 months – 60 years who present with anemia.
2. Patients willing to participate in the study.

Exclusion criteria

1. Patients aged <6 months ad >60 years.
2. Patients with multiple blood transfusions within three months of screening for SCD.
3. Patients who are not willing to participate in the study.

Method

Potential participants will be selected on the basis of their chief complaints such as clinical anemia, any bone pain, family history along with complete blood count report and peripheral blood examinations suggesting hemolytic anemia. After that, we will do the sickling test and the correlation with peripheral blood examination will be done. If the sickling test is positive, then we will do Hb electrophoresis which will confirm type of hemolytic anemia on the basis of presence of abnormal hemoglobin such as HbS. This will further confirm the type of hemolytic anemia as sickle cell anemia (SCA). Informed consent will be obtained prior to participation in the study. The patient will be comfortably seated and a blood sample will be taken under aseptic precautions. 5ml blood will be drawn into a dipotassium ethylenediaminetetraacetic acid (K2EDTA) tube through a standard central cubital vein, for testing of SCD.

All patients will be examined further for

1. Sickling test
2. Peripheral Blood smear
3. Hemoglobin electrophoresis

Sickling test

The sickling test’s basic method depends on microscopic observations of red blood cells (RBC) that are exposed to low oxygen tension or start to swell. An equal volume of EDTA blood will be mixed with 2% sodium metabisulfite. A drop of blood and a mixture of reagents will be taken on a glass slide. A cover slip will be placed over it, and the four corners will be sealed with wax and the slide will be observed under a microscope for presence of any sickle shaped red blood cells if any (40x) after 2, 4, and 24 hrs.

Peripheral blood smear

The peripheral blood smear (PBF) examines the blood cell’s morphology and evaluates any microscopic alterations, which can offer useful information that helps in the diagnosis of SCD.

The method is as follows: the smear will form a thin film of blood. Allow it to air dry, and then place it in a staining rack. Leishman’s stain will be placed dropwise on the slide and left for 2 minutes. This allows the determination of PBF in methyl alcohol. Add a double volume of buffered water dropwise to the slide and let stand for 8 minutes and then rinse in water for 1 to 2 minutes. Air dry and examine under the oil immersion lens of a microscope.

Hemoglobin electrophoresis

Equal parts of distilled water and packed cells will be combined and then the mixture will be centrifuged. As a sample, this hemolysate will be used. Medium 2% agarose gel pH 8.6 with Tris/EDTA/borate (TEB) buffer will be used. The basic idea behind this method is based on the fact that proteins can be positively or negatively charged based on the amino acids that are normally charged.

The positively charged proteins will move to the cathode when an electric field is applied to a solution containing protein molecules, and vice versa. Different hemoglobin will separate and migrate at different rates depending on their charges, size, and shape. Following this, bands are stained and their color concentration is scanned in comparison to known controls.

Bias

All necessary measures to control bias at all levels will be taken. As it is an observational study, all the samples which are routinely received in the section of hematology will be taken into consideration for comparison between the Hb electrophoresis technique analogous to the routine sickling test. Institutional supply of EDTA samples and chemical constituents present in a buffer from different companies make minor modifications, essential part of Hb electrophoresis protocol. Diagnostic search influences the finding of electrophoresis sickling band.

Sample size

Daniel’s formula for sample size

n = \frac{Z^{2} p (1 - p)}{d^{2}}

Where,

Z∝2 statistics for a level of confidence (for the level of confidence of 95%, which is conventional, the Z value is 1.96) = 1.960

p = prevalence of sickle cell anemia = 5 % = 0.05

D = desired error of margin = 6% = 0.06

n is the no. of patients is 72.99

\begin{array}{c} n = \frac{{1.96}^{2 *} {0.05}^{*} (1 - 0.05)}{{0.05}^{2}} \end{array}

$n = 72.99$

$= 75 patients needed in the study .$

Formula reference: Daniel et al. (1977).¹⁸

Outcome

Primary outcomes

We will observe any sickle shaped red blood cells under the microscope which can be easily seen in case of sickle cell disease (SCD) patients, but gold standard for diagnosis of SCD is always hemoglobin electrophoresis which shows the percentage of HbS hemoglobin on its report.

Expected outcome

To evaluate the sensitivity and specificity of the sickling test, peripheral blood smear, and hemoglobin electrophoresis for SCD screening.

Data analytics and statistical plan

SPSS version 23.0 will be used to conduct the statistical analysis. A p value of 0.05 will be regarded as significant. The mean distribution of inhibition zones will be documented as part of the descriptive analysis. Patient and repeated measures will be analysed using ANOVA.

Dissemination

The results will be published in an indexed journal.

Study status

The study is set to start in September 2023.

Results

The results will inform healthcare professionals and policymakers about the effectiveness of these methods and contribute to improved screening strategies for early detection and management of sickle cell disease.

Discussion

The screening techniques include the sickling and peripheral blood film examination. We will collect the blood samples and we will evaluate these for validity of the sickling test and peripheral blood film. Clinical anemia is present in patients ranging from age 6 months to 60 years till now.

Sickle cell disease is the most common a single-gene disorder correlated with recurrent episodes of intense ailment and myeloproliferative. Herrick originally recognised SCD in a Chicago-based West Indian patient with anemia and heart disturbances in 1910.³ After he published his paper describing the sickle-shaped haemocyte, Pauling determined the deformity of hemoglobin in SCD and invented the term “molecular disease” in 1949.¹⁹

India forms a major part of all worldwide cases of sickle cell anemia (SCA).¹

A study conducted by A. L. Okwietal (2010),‘The Reliability of Sickling and Solubility Tests and Peripheral Blood Film Methods for Sickle Cell Disease Screening at District Health Centers in Uganda’, reported that sensitivity estimations for the tests of solubility and sickling were 65.0% and 45.0%, respectively, while the peripheral film had a 35.0% sensitivity. There were 95.6%, 90.0%, and 96.7% specificities for sickling, solubility, and peripheral film, respectively. Sickling received a diagnosis of 92.5% accuracy, 85.5% solubility, and 90.5% of peripheral film. Cohen’s Kappa for sickling was 0.6, solubility was 0.3, and peripheral film was 0.4. Turnaround time for the sickling test was 38 minutes, solubility was 70 minutes, and peripheral was 44 minutes.¹⁵

Limitations

False-positive results occur when the test indicates the presence of SCD when it is not actually present, leading to unnecessary concern and follow-up testing. False-negative results occur when the test fails to identify individuals with SCD, delaying necessary interventions and treatment.

The reliability of the sickling test and peripheral blood film method may vary with the age of the individual being tested. Some infants with SCD may not show symptoms or positive test results immediately after birth, leading to potential delays in diagnosis. The accuracy of both methods is highly dependent on the technical expertise of the personnel performing the tests. Improper sample collection, preparation, or interpretation can lead to inaccurate results. Distinguishing between sickle cell trait (SCT) and SCD is essential, as individuals with SCT are carriers of the gene but typically do not experience symptoms or complications. Both screening methods may not always differentiate between SCT and SCD.

The quality of the blood sample used for testing can impact the accuracy of the results. Factors such as hemolysis or contamination can affect the reliability of the tests.

Interpretation

Sickling method confirms abnormal hemoglobin presence, while peripheral blood smear reveals sickle-shaped red blood cells, aiding the diagnosis of sickle cell disease and assessing disease severity.

Generalisability

Peripheral smear and sickling evaluation offer a reliable and widely applicable method for diagnosing hemoglobin disorders in the general population.

Data availability

No data are associated with this article.

Reporting guidelines

Zenodo: STROBE checklist for ‘Evaluating the reliability of the sickling test and peripheral blood film method for screening of sickle cell disease: a study protocol’. https://doi.org/10.5281/zenodo.8199129

Acknowledgments

We would like to thank our institute and colleagues. We acknowledge the support of Sonali Jakkulwar in the conceptualisation and preparation of this manuscript.

References

1. Gorakshakar AC: Epidemiology of sickle hemoglobin in India. In Proceeding of the National Symposium on Tribal Health 2006 Oct 19. (pp. 103–108).
2. Disorders of hemoglobin structures and synthesis. de Gruchy’sClinicalHaematology in Medical Practice. 5th edition.Oxford University Press; 1990; 171–87.
3. Herrick JB: Peculiar elongated and sickle-shaped red blood corpuscles in a case of severe anemia. 1910. Yale J. Biol. Med. 2001 May; 74(3): 179–184. PubMed Abstract
4. Kato GJ, Piel FB, Reid CD, et al.: Sickle cell disease. Nat. Rev. Dis. Primers. 2018 Mar 15; 4(1): 1–22. Publisher Full Text
5. Nagel RL, Johnson J, Bookchin RM, et al.: Β-Chain contact sites in the hemoglobin S polymer. Nature. 1980 Feb 28; 283(5750): 832–834. PubMed Abstract | Publisher Full Text
6. Benesch RE, Kwong S, Benesch R: The effects of α chain mutations cis and trans to the β 6 mutation on the polymerization of sickle cell hemoglobin. Nature. 1982 Sep 16; 299(5880): 231–234. PubMed Abstract | Publisher Full Text
7. Bee R, Srivastav AK: TO DETECT HEMOGLOBINOPATHIES BY DOING HEMOGLOBIN ELECTROPHORESIS IN MICROCYTIC HYPOCHROMIC ANEMIA. European Journal of Molecular & Clinical Medicine. 2021; 7(09): 2020.
8. Pomburg T, Ha SSJ, Wells IL: Sickle Cell Anemia, a Molecular Disease. Science. 1949; 110: 543–548. Publisher Full Text
9. Krishna OR, Bapanpally N, Fatima SS, et al.: Quantification of Sickle Cells in the Peripheral Smear as a Marker of Disease Severity in Sickle Cell Disease in Paediatric Patients. J. Med. Sci. 2023 Jan; 9(1): 104–109. Publisher Full Text
10. Lubeck D, Agodoa I, Bhakta N, et al.: Estimated life expectancy and income of patients with sickle cell disease compared with those without sickle cell disease. JAMA Netw. Open. 2019 Nov 1; 2(11): e1915374. PubMed Abstract | Publisher Full Text | Free Full Text
11. Jadhav U, Babu R, Ghewade B, et al.: Acute chest syndrome, a distinctive manifestation in sickle cell disease–A case study. J. Datta Meghe Inst. Med. Sci. Univ. 2020 Jul 1; 15(3): 492–494. Publisher Full Text
12. Varma A, Vagha J, Agrawal A, et al.: Sociodemographic determinants in prevalence of anemia in adolescents of rural area of Maharashtra. J. Datta Meghe Inst. Med. Sci. Univ. 2020 Apr 1; 15(2): 209–214. Publisher Full Text
13. Mithra P, Khatib MN, Sinha AP, et al.: Interventions for addressing anemia among children and adolescents: an overview of systematic reviews. Front. Pediatr. 2021 Feb 16; 8: 549549. PubMed Abstract | Publisher Full Text | Free Full Text
14. Agrawal S, Deshmukh P, Deshmukh P, et al.: Anesthetic Management of Patients with Sickle Cell Disease Posted for Bipolar Prosthesis. Indian. J. Forensic Med. Toxicol. 2020 Oct 1; 14(4).
15. Okwi AL, Byarugaba W, Parkes A, et al.: The reliability of sickling and solubility tests and peripheral blood film method for sickle cell disease screening at district health centers in Uganda. Clinics in Mother and Child Health. 2010 Oct 6; 7(1): 1–5. Publisher Full Text
16. Arishi WA, Alhadrami HA, Zourob M: Techniques for the detection of sickle cell disease: a review. Micromachines. 2021 May 5; 12(5): 519. PubMed Abstract | Publisher Full Text | Free Full Text
17. Dada EG, Oyewola DO, Joseph SB: Deep convolutional neural network model for detection of sickle cell anemia in peripheral blood images. Communication in Physical Sciences. 2022 Mar 8; 8(1).
18. Daniel et al. (1977).
19. Pauling L, Itano HA, Singer SJ, et al.: Sickle cell anemia, a molecular disease. Science. 1949; 110(1): 543–548. Publisher Full Text

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¹ Pathology, Datta Meghe Institute of Higher Education and Research, Sawangi (M), Wardha India, Wardha, Maharashtra, 442001, India
² Pathology, Datta Megha Institute of Higher Education and Research Sawangi (M) Wardha. India, Wardha, Maharashtra, 442001, India
³ Pathology, Datta Meghe Institute of Higher Education and Research, Sawangi (M), Wardha. India, Wardha, Maharashtra, 442001, India

Tejaswini Bawane
Roles: Investigation, Methodology, Validation, Visualization, Writing – Original Draft Preparation

Sahitya Vodithala
Roles: Investigation, Methodology, Visualization

Competing interests

No competing interests were disclosed.

Grant information

The author(s) declared that no grants were involved in supporting this work.

Article Versions (1)

version 1

Published: 11 Sep 2023, 12:1124

https://doi.org/10.12688/f1000research.139432.1

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© 2023 Bawane T and Vodithala S. This is an open access article distributed under the terms of the Creative Commons Attribution License, which permits unrestricted use, distribution, and reproduction in any medium, provided the original work is properly cited.

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Bawane T and Vodithala S. Evaluating the reliability of the sickling test and peripheral blood film method for screening of sickle cell disease: a study protocol [version 1; peer review: 1 approved with reservations]. F1000Research 2023, 12:1124 (https://doi.org/10.12688/f1000research.139432.1)

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Reviewer Report 13 May 2024

Brian D. Adkins, University of Texas Southwestern, Dallas, Texas, USA

Approved with Reservations

https://doi.org/10.5256/f1000research.152707.r242309

This is an interesting protocol describing Hemoglobin S screening. There are several points which require attention. Please see detailed explanation below.

Introduction:
Please focus content of introduction on what is being investigated in this study. Please ... Continue reading

This is an interesting protocol describing Hemoglobin S screening. There are several points which require attention. Please see detailed explanation below.

Introduction:
Please focus content of introduction on what is being investigated in this study. Please remove general paragraph about anemia, as the specialized audience will be aware of the described information.
Line 1 – please refer to hemoglobin as an oxygen containing molecule that gives RBCs their red color. Calling hemoglobin, a pigment is too reductive. Also please use a PubMed indexed reference.
Rationale
Line 1 – this repeats the information contained in paragraph 2 of introduction. Please incorporate most of this data into the introduction and focus this section on the rationale.
Third sentence first paragraph: please include examples of false positives/negatives from package insert.
“There aren’t many skilled pathologists to choose from” too colloquial, remove/reword please.
Finally, would point out that some countries, such as the USA have routine newborn screening for SCD and other hemoglobinopathies.
Discussion
“Clinical anemia is present in patients ranging from age 6 months to 60 years till now” not true, please remove.
Generalizability
We don’t know if these tests are generalizable because the study has not been completed. Please reword to reflect that.

Is the rationale for, and objectives of, the study clearly described?

Yes
Is the study design appropriate for the research question?

Yes
Are sufficient details of the methods provided to allow replication by others?

Yes
Are the datasets clearly presented in a useable and accessible format?

Not applicable

Competing Interests: No competing interests were disclosed.

Reviewer Expertise: Clinical transfusion medicine, blood banking, academic pathology, DEI, hematopathology

I confirm that I have read this submission and believe that I have an appropriate level of expertise to confirm that it is of an acceptable scientific standard, however I have significant reservations, as outlined above.

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13 May 2024 | for Version 1

Brian D. Adkins, University of Texas Southwestern, Dallas, Texas, USA

6 Views Cite this report Responses(0)

Approved With Reservations

This is an interesting protocol describing Hemoglobin S screening. There are several points which require attention. Please see detailed explanation below.

Introduction:
Please focus content of introduction on what is being investigated in this study. Please remove general paragraph about anemia, as the specialized audience will be aware of the described information.
Line 1 – please refer to hemoglobin as an oxygen containing molecule that gives RBCs their red color. Calling hemoglobin, a pigment is too reductive. Also please use a PubMed indexed reference.
Rationale
Line 1 – this repeats the information contained in paragraph 2 of introduction. Please incorporate most of this data into the introduction and focus this section on the rationale.
Third sentence first paragraph: please include examples of false positives/negatives from package insert.
“There aren’t many skilled pathologists to choose from” too colloquial, remove/reword please.
Finally, would point out that some countries, such as the USA have routine newborn screening for SCD and other hemoglobinopathies.
Discussion
“Clinical anemia is present in patients ranging from age 6 months to 60 years till now” not true, please remove.
Generalizability
We don’t know if these tests are generalizable because the study has not been completed. Please reword to reflect that.

Is the rationale for, and objectives of, the study clearly described?

Yes
Is the study design appropriate for the research question?

Yes
Are sufficient details of the methods provided to allow replication by others?

Yes
Are the datasets clearly presented in a useable and accessible format?

Not applicable

Competing Interests

No competing interests were disclosed.

Reviewer Expertise

Clinical transfusion medicine, blood banking, academic pathology, DEI, hematopathology

I confirm that I have read this submission and believe that I have an appropriate level of expertise to confirm that it is of an acceptable scientific standard, however I have significant reservations, as outlined above.

Respond to this report

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[1] 1. Gorakshakar AC: Epidemiology of sickle hemoglobin in India. In Proceeding of the National Symposium on Tribal Health 2006 Oct 19. (pp. 103–108).

[2] 2. Disorders of hemoglobin structures and synthesis. de Gruchy’sClinicalHaematology in Medical Practice. 5th edition.Oxford University Press; 1990; 171–87.

[3] 3. Herrick JB: Peculiar elongated and sickle-shaped red blood corpuscles in a case of severe anemia. 1910. Yale J. Biol. Med. 2001 May; 74(3): 179–184. PubMed Abstract

[4] 4. Kato GJ, Piel FB, Reid CD, et al.: Sickle cell disease. Nat. Rev. Dis. Primers. 2018 Mar 15; 4(1): 1–22. Publisher Full Text

[5] 5. Nagel RL, Johnson J, Bookchin RM, et al.: Β-Chain contact sites in the hemoglobin S polymer. Nature. 1980 Feb 28; 283(5750): 832–834. PubMed Abstract | Publisher Full Text

[6] 6. Benesch RE, Kwong S, Benesch R: The effects of α chain mutations cis and trans to the β 6 mutation on the polymerization of sickle cell hemoglobin. Nature. 1982 Sep 16; 299(5880): 231–234. PubMed Abstract | Publisher Full Text

[7] 7. Bee R, Srivastav AK: TO DETECT HEMOGLOBINOPATHIES BY DOING HEMOGLOBIN ELECTROPHORESIS IN MICROCYTIC HYPOCHROMIC ANEMIA. European Journal of Molecular & Clinical Medicine. 2021; 7(09): 2020.

[8] 8. Pomburg T, Ha SSJ, Wells IL: Sickle Cell Anemia, a Molecular Disease. Science. 1949; 110: 543–548. Publisher Full Text

[9] 9. Krishna OR, Bapanpally N, Fatima SS, et al.: Quantification of Sickle Cells in the Peripheral Smear as a Marker of Disease Severity in Sickle Cell Disease in Paediatric Patients. J. Med. Sci. 2023 Jan; 9(1): 104–109. Publisher Full Text

[10] 10. Lubeck D, Agodoa I, Bhakta N, et al.: Estimated life expectancy and income of patients with sickle cell disease compared with those without sickle cell disease. JAMA Netw. Open. 2019 Nov 1; 2(11): e1915374. PubMed Abstract | Publisher Full Text | Free Full Text

[11] 11. Jadhav U, Babu R, Ghewade B, et al.: Acute chest syndrome, a distinctive manifestation in sickle cell disease–A case study. J. Datta Meghe Inst. Med. Sci. Univ. 2020 Jul 1; 15(3): 492–494. Publisher Full Text

[12] 12. Varma A, Vagha J, Agrawal A, et al.: Sociodemographic determinants in prevalence of anemia in adolescents of rural area of Maharashtra. J. Datta Meghe Inst. Med. Sci. Univ. 2020 Apr 1; 15(2): 209–214. Publisher Full Text

[13] 13. Mithra P, Khatib MN, Sinha AP, et al.: Interventions for addressing anemia among children and adolescents: an overview of systematic reviews. Front. Pediatr. 2021 Feb 16; 8: 549549. PubMed Abstract | Publisher Full Text | Free Full Text

[14] 14. Agrawal S, Deshmukh P, Deshmukh P, et al.: Anesthetic Management of Patients with Sickle Cell Disease Posted for Bipolar Prosthesis. Indian. J. Forensic Med. Toxicol. 2020 Oct 1; 14(4).

[15] 15. Okwi AL, Byarugaba W, Parkes A, et al.: The reliability of sickling and solubility tests and peripheral blood film method for sickle cell disease screening at district health centers in Uganda. Clinics in Mother and Child Health. 2010 Oct 6; 7(1): 1–5. Publisher Full Text

[16] 16. Arishi WA, Alhadrami HA, Zourob M: Techniques for the detection of sickle cell disease: a review. Micromachines. 2021 May 5; 12(5): 519. PubMed Abstract | Publisher Full Text | Free Full Text

[17] 17. Dada EG, Oyewola DO, Joseph SB: Deep convolutional neural network model for detection of sickle cell anemia in peripheral blood images. Communication in Physical Sciences. 2022 Mar 8; 8(1).

[18] 18. Daniel et al. (1977).

[19] 19. Pauling L, Itano HA, Singer SJ, et al.: Sickle cell anemia, a molecular disease. Science. 1949; 110(1): 543–548. Publisher Full Text

Evaluating the reliability of the sickling test and peripheral blood film method for screening of sickle cell disease: a study protocol

Abstract

Keywords

Introduction

Rationale

Protocol

Aim

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Outcome

Data analytics and statistical plan

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Study status

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Generalisability

Data availability

Reporting guidelines

Acknowledgments

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