last row of a gridview can empty validation is not needed .

Your question is a bit confusing for me. Can you provide additional information, including the problem you're encountering and what result you're expecting?

If you want to skip validation on last row of the gridview then:

for (int i = 0; i < gvAcademicQualification.Rows.Count; i++)
{
    // Skip validation for the last row
    if (i == gvAcademicQualification.Rows.Count - 1)
    {
        continue;
    }

    GridViewRow row = gvAcademicQualification.Rows[i];

    TextBox txtStream = (TextBox)row.FindControl("txtStream");
    TextBox txtInstituteName = (TextBox)row.FindControl("txtInstituteName");
    TextBox txtGrade = (TextBox)row.FindControl("txtGrade");
    TextBox txtYearOfPassing = (TextBox)row.FindControl("txtYearOfPassing");

    RequiredFieldValidator rtxtStream = (RequiredFieldValidator)row.FindControl("rtxtStream");
    RequiredFieldValidator rtxtInstituteName = (RequiredFieldValidator)row.FindControl("rtxtInstituteName");
    RequiredFieldValidator rtxtGrade = (RequiredFieldValidator)row.FindControl("rtxtGrade");
    RequiredFieldValidator rtxtYearOfPassing = (RequiredFieldValidator)row.FindControl("rtxtYearOfPassing");

    rtxtStream.Visible = string.IsNullOrEmpty(txtStream.Text);
    rtxtInstituteName.Visible = string.IsNullOrEmpty(txtInstituteName.Text);
    rtxtGrade.Visible = string.IsNullOrEmpty(txtGrade.Text);
    rtxtYearOfPassing.Visible = string.IsNullOrEmpty(txtYearOfPassing.Text);
}

If you want to perform validation for the first, second, and third rows:

for (int i = 0; i < gvAcademicQualification.Rows.Count; i++)
{
    if (i >= 3)
    {
        continue;
    }

    GridViewRow row = gvAcademicQualification.Rows[i];

    TextBox txtStream = (TextBox)row.FindControl("txtStream");
    TextBox txtInstituteName = (TextBox)row.FindControl("txtInstituteName");
    TextBox txtGrade = (TextBox)row.FindControl("txtGrade");
    TextBox txtYearOfPassing = (TextBox)row.FindControl("txtYearOfPassing");

    RequiredFieldValidator rtxtStream = (RequiredFieldValidator)row.FindControl("rtxtStream");
    RequiredFieldValidator rtxtInstituteName = (RequiredFieldValidator)row.FindControl("rtxtInstituteName");
    RequiredFieldValidator rtxtGrade = (RequiredFieldValidator)row.FindControl("rtxtGrade");
    RequiredFieldValidator rtxtYearOfPassing = (RequiredFieldValidator)row.FindControl("rtxtYearOfPassing");

    rtxtStream.Visible = string.IsNullOrEmpty(txtStream.Text);
    rtxtInstituteName.Visible = string.IsNullOrEmpty(txtInstituteName.Text);
    rtxtGrade.Visible = string.IsNullOrEmpty(txtGrade.Text);
    rtxtYearOfPassing.Visible = string.IsNullOrEmpty(txtYearOfPassing.Text);
}

Please test this

for (int i = 0; i < gvAcademicQualification.Rows.Count; i++)
{
    GridViewRow row = gvAcademicQualification.Rows[i];

    if (i == 3)
    {
        TextBox txtStream = (TextBox)row.FindControl("txtStream");
        TextBox txtInstituteName = (TextBox)row.FindControl("txtInstituteName");
        TextBox txtGrade = (TextBox)row.FindControl("txtGrade");
        TextBox txtYearOfPassing = (TextBox)row.FindControl("txtYearOfPassing");

        // Use the correct RequiredFieldValidators for each TextBox
        RequiredFieldValidator rtxtStream = row.FindControl("rtxtStream") as RequiredFieldValidator;
        RequiredFieldValidator rtxtInstituteName = row.FindControl("rtxtInstituteName") as RequiredFieldValidator;
        RequiredFieldValidator rtxtGrade = row.FindControl("rtxtGrade") as RequiredFieldValidator;
        RequiredFieldValidator rtxtYearOfPassing = row.FindControl("rtxtYearOfPassing") as RequiredFieldValidator;

        // Perform validation for the desired fields
        if (string.IsNullOrEmpty(txtStream.Text))
        {
            // Handle the validation error for Stream field
            rtxtStream.Visible = true;
        }

        if (string.IsNullOrEmpty(txtInstituteName.Text))
        {
            // Handle the validation error for Institute Name field
            rtxtInstituteName.Visible = true;
        }

        if (string.IsNullOrEmpty(txtGrade.Text))
        {
            // Handle the validation error for Grade field
            rtxtGrade.Visible = true;
        }

        if (string.IsNullOrEmpty(txtYearOfPassing.Text))
        {
            // Handle the validation error for Year of Passing field
            rtxtYearOfPassing.Visible = true;
        }
    }
}